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Cited by 92Protein Kinase Regulation and GTPase SignalingProtein Structure and DynamicsProtein Tyrosine Phosphatases

Assessment of mutation probabilities of KRAS G12 missense mutants and their long-timescale dynamics by atomistic molecular simulations and Markov state modeling

Open access
Pantsar, Tatu, Rissanen, Sami, Dauch, Daniel, Laitinen, Tuomo, Vattulainen, Ilpo, Poso, Antti·2018·PLOS Computational Biology
DOI 10.1371/journal.pcbi.1006458PMC PMC6147662
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Abstract

A mutated KRAS protein is frequently observed in human cancers. Traditionally, the oncogenic properties of KRAS missense mutants at position 12 (G12X) have been considered as equal. Here, by assessing the probabilities of occurrence of all KRAS G12X mutations and KRAS dynamics we show that this assumption does not hold true. Instead, our findings revealed an outstanding mutational bias. We conducted a thorough mutational analysis of KRAS G12X mutations and assessed to what extent the observed mutation frequencies follow a random distribution. Unique tissue-specific frequencies are displayed with specific mutations, especially with G12R, which cannot be explained by random probabilities. To clarify the underlying causes for the nonrandom probabilities, we conducted extensive atomistic molecular dynamics simulations (170 μs) to study the differences of G12X mutations on a molecular level. The simulations revealed an allosteric hydrophobic signaling network in KRAS, and that protein dynamics is altered among the G12X mutants and as such differs from the wild-type and is mutation-specific. The shift in long-timescale conformational dynamics was confirmed with Markov state modeling. A G12X mutation was found to modify KRAS dynamics in an allosteric way, which is especially manifested in the switch regions that are responsible for the effector protein binding. The findings provide a basis to understand better the oncogenic properties of KRAS G12X mutants and the consequences of the observed nonrandom frequencies of specific G12X mutations.

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Written from the paper’s full text.

TL;DR

KRAS G12X mutation frequencies are nonrandom and tissue-specific, and atomistic simulations show each G12X mutant has distinct long-timescale dynamics relative to wild-type. The authors link these mutation-specific dynamic changes to an allosteric hydrophobic hub network and altered switch-region conformations.

Key findings

  • In 32,654 tumor samples with KRAS G12X missense mutations, the authors analyzed eight tissues with >10% occurrence, comprising 31,251 samples (95%); the large intestine (18,174), lung (5,640), and pancreas (5,528) dominated. Overall frequencies were G12D 42%, G12V 28%, G12C 14%, with G12A/G12R/G12S less common; G12R was 13% in pancreas but <2% in small intestine, and G12C was most abundant in lung.
  • Mutation position was biased: 76.6% were c.35G>X (G12A, G12D, G12V) and 23.4% were c.34G>X (G12C, G12R, G12S). Base substitutions were G>A 47.4%, G>T 42.1%, G>C 10.5%; c.35 preference was >7x for G>A, 1.75x for G>C, and 2.03x for G>T, with tissue exceptions such as pancreas G>C occurring nine times more often at c.34 and lung G>T preferring c.34 >1.5x.
  • A total of 170 μs (85 × 2 μs) of atomistic MD simulations of wild-type KRAS and all G12X mutants with GDP/GTP revealed mutation-specific changes in long-timescale dynamics, especially in switch-I and switch-II, despite no differences at residue 12 itself.
  • PCA, interaction-network analysis, and MSMs identified an 11-hub hydrophobic allosteric network (V14, M72, F78, L79, F90, I100, V114, A146, A155, F156, L159); MSM found seven metastable states, with states VI and VII populated 77–87%, G12R uniquely occupying state I (6%), G12V state III (6%), and G12V shifted toward perpendicular switch states (12% vs ~1% for others).

Methods

  • Mutation frequencies were compiled from COSMIC v79, including only single-nucleotide substitutions and tissues with >10% KRAS G12X occurrence; Fisher’s exact test assessed differences.
  • MD simulations used GROMACS 4.6 with OPLS-AA, starting from GDP-bound wild-type KRAS (PDB 4OBE), with GDP or GTP, TIP3P water, 140 mM K+/Cl−, NpT at 310 K and 1 atm, 2 fs time step, and 5–10 independent 2 μs replicas per system (total 170 μs).
  • Analyses included RMSF, backbone PCA using the last 1.7 μs after discarding 300 ns, PyInteraph interaction-network analysis, and PyEMMA 2 MSMs built from selected inter-residue distances with TICA (40 ns lag), k-means clustering, and PCCA++ to define seven metastable states.

Limitations

  • The authors state that the HVR and cell membrane were absent from simulations, so they cannot deduce whether mutational effects attenuate or amplify through these factors; membrane-related effects remain for follow-up studies.
  • The authors state that MSM analysis focused on wild-type and the most important mutants G12D, G12R, and G12V because MSMs require extended simulation data, and that interaction-network analysis reports interaction frequencies rather than exact interaction characteristics.
  • Evident from the text: mutation-frequency analysis was restricted to single-nucleotide substitutions and tissues with >10% occurrence, and the simulations showed no differences at residue 12 or its vicinity, with individual replica variation sometimes as large as differences between systems.

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Integrity signals

Registry

Crossref · 10.1371/journal.pcbi.1006458

Cites

86 of 86 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

0 of 88 cited sources checked · 0 supported · 0 flagged

  • Findingown data

    KRAS G12X mutation frequencies display an outstanding mutational bias.

    “Intriguingly, an outstanding mutational bias emerged from the data.”
    Quote in the paperSubstantial claim
  • Findingown data

    The G12X mutation is far from the binding interface.

    “Given this, and since the G12X mutation is far from the binding interface (S1B Fig)”
    Quote in the paperSubstantial claim
  • Findingown data

    KRAS contains an allosteric hydrophobic signaling network.

    “The simulations revealed an allosteric hydrophobic signaling network in KRAS”
    Quote in the paperSubstantial claim
  • Findingown data

    The shift in long-timescale conformational dynamics of KRAS G12X mutants was confirmed with Markov state modeling.

    “The shift in long-timescale conformational dynamics was confirmed with Markov state modeling.”
    Quote in the paperSubstantial claim
  • Findingcitation

    A total of 32,654 tumor samples identified with a KRAS G12X missense mutation were found from the COSMIC database.

    “A total of 32,654 tumor samples identified with a KRAS G12X missense mutation were found from the database.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [2] SA Forbes, 2017, COSMIC: somatic cancer genetics at high-resolution, Nucleic Acids Res source

      Not checked in the free run.

  • May combine two claims. 62
  • Classified as probably not a substantial claim. 47

  • May combine two claims.

    Findingown data

    The results provided compelling evidence that mutations alter the dynamics of KRAS, that the alteration is mutation specific, displays allosteric characteristics, and is manifested especially in the effector protein-binding interface.

    “The results provided compelling evidence that mutations alter the dynamics of KRAS, that the alteration is mutation specific, displays allosteric characteristics, and that the alteration is manifested especially in the effector protein-binding interface.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Eight tissue types with >10% KRAS G12X mutations comprised 31,251 positive samples, 95% of all KRAS G12X mutations in the database.

    “This status is displayed in eight tissue types, which in total comprised 31,251 positive samples (95% of all KRAS G12X mutations in the database).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    KRAS protein dynamics is altered among G12X mutants and differs from wild-type in a mutation-specific manner.

    “protein dynamics is altered among the G12X mutants and as such differs from the wild-type and is mutation-specific.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    KRAS G12X mutation frequencies vary by tissue, with specific mutations such as G12R showing tissue-specific frequencies.

    “Unique tissue-specific frequencies are displayed with specific mutations, especially with G12R, which cannot be explained by random probabilities.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    The large intestine (18,174), lung (5,640), and pancreas (5,528) had numerous positive samples, whereas biliary tract, endometrium, ovary, peritoneum, and small intestine comprised altogether only 2,085 positive samples.

    “The large intestine (18,174), the lung (5,640), and the pancreas (5,528) were observed to have numerous positive samples, whereas the biliary tract, the endometrium, the ovary, the peritoneum, and the small intestine comprised altogether only 2,085 positive samples.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    The switch regions in HRAS and KRAS are identical, so their expected binding mode to their effectors is alike.

    “Given that the switch regions in HRAS and KRAS are identical (S1C Fig), their expected binding mode to their effectors is alike.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingreasoning

    G12D mutation is consistent with random expectation, as it occurs very often in all tissues.

    “G12D mutation is consistent with this view, as it occurs very often in all tissues.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Score plots for principal components PC1-3 highlight dissimilarities between wild-type KRAS and the mutants, as well as between GDP- and GTP-bound proteins.

    “The results highlight dissimilarities between the wild-type KRAS and the mutants, as well as between GDP- and GTP-bound proteins.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingcitation

    The G12R mutation is observed in the pancreas with a probability of 13%, while in the small intestine it appears in less than 2% of cases.

    “For instance, the G12R mutation is observed in the pancreas with a probability of 13%, while in the small intestine it appears in less than 2% of the cases.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    The lung differs from other tissues with a higher G>T fraction and a diminished proportion of G>A mutations (P < 0.001).

    “Particularly the lung differs from other tissues with a higher G>T fraction and a diminished proportion of G>A mutations (P < 0.001).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Overall, G12D (42%), G12V (28%), and G12C (14%) mutations are very common, whereas G12A, G12R, and G12S are less popular.

    “Overall, G12D (42%), G12V (28%), and G12C (14%) mutations are very common, whereas G12A, G12R, and G12S are less popular.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    From the hub A155, both G12A and G12S lack interactions to V114, L79, and I142, but have elevated interaction to F156, and G12A has an additional interaction to V152.

    “From the hub A155, both lack the interactions to V114, L79, and I142, but instead they have an elevated interaction to F156, and the G12A has an additional interaction to V152 (S11C Fig).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    The predominating mutations are G12D and G12V, with the lung being an exception where G12C is the most abundant mutation.

    “The predominating mutations are G12D and G12V, the lung being an exception with G12C standing as the most abundant mutation.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    The positional mutation preference for c.35G>X is highest with a G>A mutation (>7x), whereas G>C or G>T exhibit nearly twofold preference, 1.75- and 2.03-fold, respectively.

    “The positional mutation preference for c.35G>X seems to be the highest with a G>A mutation (>7x), whereas G>C or G>T exhibit nearly twofold preference, 1.75- and 2.03-fold, respectively (Fig 1M).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Individual PCA analyses highlight variation in switch region movements among systems and show that specific systems display more dynamical behavior in the α3-helix, hairpin loop between β2- and β3-sheets, α4-helix, loop between β5-sheet and α4-helix, and SAK-motif regions.

    “The individual PCA analyses highlight not just variation in the switch region movements among the systems, but they also show that specific systems display more dynamical behavior in the α3-helix, hairpin loop between the β2- and β3-sheets, the α4-helix, the loop between β5-sheet and α4-helix, and in the SAK-motif (residues 145–147) regions.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    The pancreas (>28x), peritoneum (>37x), and ovary (>41x) exhibit the most prominent preference for the G>A mutation in c.35.

    “Above all, the pancreas (>28x), the peritoneum (>37x) and the ovary (>41x) exhibit the most prominent preference for the G>A mutation in c.35.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    PC1 of GTP-bound systems displayed movement only in the switch regions, whereas PC1 of GDP-bound systems exhibited additional movement in the α3-helix.

    “PC1 of the GTP-bound systems displayed movement only in the switch regions, whereas PC1 of the GDP-bound systems exhibited additional movement also in the α3-helix.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    76.6% of G12X mutations are c.35G>X mutations (G12A, G12D, G12V) and only 23.4% are c.34G>X mutations (G12C, G12R, G12S).

    “Interestingly, 76.6% of the G12X mutations are c.35G>X mutations (G12A, G12D, G12V) and only 23.4% are c.34G>X mutations (G12C, G12R, G12S) (Fig 1L).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    A hydrophobic hub network in KRAS connects distant sites in the structure and is able to convey effects in an allosteric way.

    “Interestingly, we identified a hydrophobic hub network in KRAS that indeed connects the distant sites in the structure and is able to convey these effects in an allosteric way (Fig 4).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    G>A and G>T mutations (47.4% and 42.1%, respectively) occur very often, while G>C mutation (10.5%) takes place more seldom.

    “G>A and G>T mutations (47.4% and 42.1%, respectively) occur very often, while the G>C mutation (10.5%) takes place more seldom.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    In the V14 hub, G12A and G12S mutants lack the interaction to A81 (<2% vs. wild-type 26.9% and 39.7%, with GDP and GTP, respectively), and display highly diminished interaction to A11 compared to wild-type KRAS.

    “For example, in the V14 hub, the G12A and G12S mutants lack the interaction to A81 (<2% vs. wild-type 26.9% and 39.7%, with GDP and GTP, respectively), and also display highly diminished interaction to A11 compared to the wild-type KRAS (S10A Fig).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    From the V114 hub, G12A and G12S mutants lack interactions to A155 and L79, but have a strengthened interaction to I142 and a totally new interaction to L113 not displayed by other systems.

    “From the V114 hub, these mutants lack interactions to A155 and L79, but instead have a strengthened interaction to I142 and a totally new interaction to L113, which is not displayed by other systems (S11A Fig).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    The conformations of G12R are clearly shifted towards the fixed switch-II states, whereas G12V is shifted away from these states towards the perpendicular states.

    “In contrast, the conformations of G12R are clearly shifted towards the fixed switch-II states, whereas G12V is shifted away from these states towards the perpendicular states.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingreasoning

    A G12X mutation alters the dynamics of the neighboring residue A11 or the whole P-loop (including both A11 and V14).

    “a G12X mutation alters the dynamics of the neighboring residue A11 or the whole P-loop (including both A11 and V14).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Metastable state V, where switch-II appears in a fixed conformation, is rarely observed with the G12V mutant (1%), whereas it is similarly represented among the other mutants and the wild-type (12-16%).

    “The moderately populated metastable state V, where switch-II appears in a fixed conformation, is rarely observed with the G12V mutant (1%), whereas it is similarly represented among the other mutants and the wild-type (12–16%).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Both D154 and R161 are located in the α5-helix, in close proximity of three hydrophobic hubs: A155, F156 and L159.

    “Both of these residues are located in the α5-helix, in the close proximity of three hydrophobic hubs: A155, F156 and L159 (Fig 4).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Metastable state I is quite unique for G12R (6%) and state III for G12V (6%), whereas the other systems are mostly absent from these two states.

    “The metastable state I is quite unique for G12R (6%) and the state III for G12V (6%), whereas the other systems are mostly absent from these two states.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    In state VII, switch-II is in a fixed conformation and switch-I is more closed compared to the first group; this group is more frequently populated by G12R.

    “In the state VII, switch-II is in a fixed conformation and switch-I is more closed compared to the first group.”
    • Also stated in: § Markov state models confirm altered dynamics in selected mutants and reveal specific metastable state distributions · plain
      “This group is more frequently populated by G12R.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    In state VI, switch-I is open and switch-II is in a mixed conformation between fixed and perpendicular; this state is clearly less populated by G12R compared to the other systems.

    “In the state VI, switch-I is open and switch-II is in a mixed conformation between the fixed and perpendicular conformations.”
    • Also stated in: § Markov state models confirm altered dynamics in selected mutants and reveal specific metastable state distributions · boosted
      “This state is clearly less populated by G12R compared to the other systems.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Group 1 (states I and V) has switch-I fully open and switch-II fixed; it is frequently occupied by G12R (24%) and mostly absent from G12V (1%).

    “The states I and V as combined form the first group, where switch-I appears to be in a fully open conformation and switch-II in a fixed conformation.”
    • Also stated in: § Markov state models confirm altered dynamics in selected mutants and reveal specific metastable state distributions · plain
      “This group is frequently occupied by G12R (24%), whereas it is mostly absent from G12V (1%).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    While wild-type does not populate the metastable states I and III at all, mutants G12R and G12V have significant population in these states (about 6%).

    “Finally, it is exceptional that while wild-type does not populate the metastable states I and III at all, there are mutants (G12R, G12V) whose population in these states is significant (about 6%).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Analysis of simulation data using PCA, interaction network analysis and MSMs revealed differences between wild-type and mutants and between mutants.

    “In our work, we unlocked this issue through the analysis of the simulation data using PCA, interaction network analysis and MSMs that indeed revealed the differences, not only between the wild-type and the mutants, but also between the mutants.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    The fourth group, where switches appear in a perpendicular conformation, is frequent with G12V (12%), whereas the other mutants rarely visit this state (1%).

    “The fourth group, where the switches appear in a perpendicular conformation, is frequent with G12V (12%), whereas the other mutants rarely visit this state (1%).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    The other mutations (G12C, G12D, G12R and G12V) are not radically different compared to wild-type in hub interactions, although some alterations are evident.

    “Surprisingly, the other mutations (G12C, G12D, G12R and G12V) are not radically different compared to the wild-type, although some alterations in the hub interactions are evident.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S4 (based).

    Findingnone given

    In specific tissues, G12X mutation frequencies exhibit an inexplicable individual bias.

    “in specific tissues G12X mutation frequencies exhibit an inexplicable individual bias.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Interpretationreasoning

    These alterations lead to increased KRAS signaling because more active GTP-bound protein is present.

    “These alterations lead to increased KRAS signaling, as there is more active GTP-bound protein present.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Premisecitation

    Smith and Ikura proposed that discrepancies in effector protein binding profiles of the mutants are due to altered switch dynamics.

    “Smith and Ikura [10] proposed that the discrepancies in the effector protein binding profiles of the mutants are due to altered switch dynamics.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [10] MJ Smith, 2014, Integrated RAS signaling defined by parallel NMR detection of effectors and regulators, Nat Chem Biol source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    RAS G12X mutants give rise to differences in preferred signaling pathway in terms of effector protein binding.

    “Importantly, not only RAS G12X mutants exhibit a discrepancy in GTP hydrolysis, but they also give rise to differences in the preferred signaling pathway (in terms of effector protein binding) [9,10].”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [9] NT Ihle, 2012, Effect of KRAS oncogene substitutions on protein behavior: Implications for signaling and clinical outcome, J Natl Cancer Inst source

      Not checked in the free run.

    • cites [10] MJ Smith, 2014, Integrated RAS signaling defined by parallel NMR detection of effectors and regulators, Nat Chem Biol source

      Not checked in the free run.

  • Classified as probably not a substantial claim.

    Premisecitation

    Mutant KRAS undergoes GDP-GTP cycling.

    “Still, the mutant KRAS undergoes GDP–GTP cycling [5].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [5] P Lito, 2016, Allele-specific inhibitors inactivate mutant KRAS G12C by a trapping mechanism, Science source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    KRAS binds GDP in its inactive state and GTP in its active state.

    “The small GTPase protein KRAS is a signal-transducing protein, which binds GDP in its inactive state and GTP in its active state [1].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [1] IR Vetter, 2001, The guanine nucleotide-binding switch in three dimensions, Science source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    KRAS G12X mutants show a discrepancy in intrinsic GTPase activity.

    “An interesting discrepancy among KRAS G12X mutants is observed in their intrinsic GTPase activity [8].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [8] JC Hunter, 2015, Biochemical and structural analysis of common cancer-associated KRAS mutations, Mol Cancer Res source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    The G12A mutation exhibits the most hindered intrinsic hydrolysis (~1% compared to wild-type), whereas G12C displays the least hindered activity (~72%).

    “The G12A mutation exhibits the most hindered intrinsic hydrolysis (~1% compared to the wild-type), whereas the G12C mutation displays the least hindered activity (~72%).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [8] JC Hunter, 2015, Biochemical and structural analysis of common cancer-associated KRAS mutations, Mol Cancer Res source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Effector proteins use a ubiquitin-like fold, a RAS-binding domain (RBD) or a RAS-association domain (RA), to bind RAS.

    “To bind RAS, the effector proteins use a ubiquitin (UB)-like fold: a RAS-binding domain (RBD) or a RAS-association domain (RA) [11,12].”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [11] CP Ponting, 1996, A novel family of RAS-binding domains, Trends Biochem Sci source

      Not checked in the free run.

    • cites [12] N Nassar, 1995, The 2.2 angstrom crystal structure of the RAS-binding domain of the serine/threonine kinase c-Raf1 in complex with Rap1A and a GTP analogue, Nature source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    The KRAS mutation is most often found at G12, in about 86% of cases.

    “The mutation is most often, in about 86% of the cases [2], found at G12.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [2] SA Forbes, 2017, COSMIC: somatic cancer genetics at high-resolution, Nucleic Acids Res source

      Not checked in the free run.

  • Classified as probably not a substantial claim.

    Premisecitation

    The basis of specific G12X mutation frequencies has remained unclear, except for the G12C transversion mutation associated with smoking in lung cancer.

    “The basis of the specific G12X mutation frequencies has remained unclear, except for the G12C transversion mutation (c.34G>T) associated with smoking in lung cancer [6,7].”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [6] IA Prior, 2012, A comprehensive survey of RAS mutations in cancer, Cancer Res source

      Not checked in the free run.

    • cites [7] S Dogan, 2012, Molecular epidemiology of EGFR and KRAS mutations in 3,026 lung adenocarcinomas: Higher susceptibility of women to smoking-related KRAS-mutant cancers, Clin Cancer Res source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    In NSCLC cell lines, KRAS G12D showed activation of PI3K and MEK signaling, while G12C and G12V exhibited activated RalGDS-pathway and diminished growth factor-dependent Akt activation.

    “This behavior was first observed in NSCLC cell lines [9], where KRAS G12D showed activation of PI3K and MEK signaling, while G12C and G12V mutants exhibited activated RalGDS-pathway and diminished growth factor-dependent Akt activation.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [9] NT Ihle, 2012, Effect of KRAS oncogene substitutions on protein behavior: Implications for signaling and clinical outcome, J Natl Cancer Inst source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    At the cellular level, isoform specificity to effector proteins is primarily determined via membrane interactions, but differences among RAS isoforms' absolute effector protein binding affinities rise from allosteric effects.

    “At the cellular level, the isoform specificity to effector proteins is primarily determined via membrane interactions [24], but the differences among RAS isoforms’ absolute effector protein binding affinities rise from allosteric effects [25].”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [24] R Nussinov, 2017, Oncogenic Ras Isoforms Signaling Specificity at the Membrane, Cancer Res source

      Not checked in the free run.

    • cites [25] CW Johnson, 2017, The small GTPases K-Ras, N-Ras, and H-Ras have distinct biochemical properties determined by allosteric effects, J Biol Chem source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Effector proteins bind to HRAS on top of its switch regions, and the binding conformation of HRAS is almost identical in all resolved complexes.

    “These effector proteins bind to HRAS on top of its switch regions: switch-I (residues 30–40) and switch-II (residues 58–72), and the binding conformation of HRAS is almost identical in all of the complexes (S1A Fig).”
    Quote in the paperSubstantial claim0 of 6 sources checked
    • cites [13] SK Fetics, 2015, Allosteric effects of the oncogenic RasQ61L mutant on Raf-RBD, Structure source

      Not checked in the free run.

    • cites [14] L Huang, 1998, Structural basis for the interaction of RAS with RalGDS, Nat Struct Biol source

      Not checked in the free run.

    • cites [15] ME Pacold, 2000, Crystal structure and functional analysis of RAS binding to its effector phosphoinositide 3-kinase gamma, Cell source

      Not checked in the free run.

    • cites [16] TD Bunney, 2006, Structural and mechanistic insights into ras association domains of phospholipase C epsilon, Mol Cell source

      Not checked in the free run.

    • cites [17] B Stieglitz, 2008, Novel type of Ras effector interaction established between tumour suppressor NORE1A and Ras switch II, EMBO J source

      Not checked in the free run.

    • cites [18] MJ Smith, 2017, Evolution of AF6-RAS association and its implications in mixed-lineage leukemia, Nat Commun source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    The oncogenic properties associated with KRAS G12X mutation are characterized by deficiency of intrinsic GTPase activity and insensitivity for GTPase-activating proteins (GAPs).

    “The oncogenic properties associated with KRAS G12X mutation are characterized by the deficiency of the intrinsic GTPase activity and the insensitivity for GTPase-activating proteins (GAPs) [3,4].”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [3] JB Gibbs, 1984, Intrinsic GTPase activity distinguishes normal and oncogenic RAS p21 molecules, Proc. Natl. Acad. Sci. USA source

      Not checked in the free run.

    • cites [4] M Trahey, 1987, A cytoplasmic protein stimulates normal N-RAS p21 GTPase, but does not affect oncogenic mutants, Science source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    A single point mutation in RAS (Q61L) has long-range effects on dynamics and alters effector protein interactions.

    “It was observed that even a single point mutation in RAS (Q61L) has long-range effects on dynamics and alters effector protein interactions [13].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [13] SK Fetics, 2015, Allosteric effects of the oncogenic RasQ61L mutant on Raf-RBD, Structure source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    The affinity of the G12C mutant to Raf is similar to that of wild-type.

    “Interestingly, the affinity of the G12C mutant is similar to that of wild-type.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [8] JC Hunter, 2015, Biochemical and structural analysis of common cancer-associated KRAS mutations, Mol Cancer Res source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    An NMR study revealed different binding preferences for mutant HRAS G12V compared to wild-type HRAS with various effector proteins, with HRAS G12V showing reduced interactions with Raf and enhanced binding with RalGDS.

    “Here, HRAS G12V showed reduced interactions with Raf and enhanced binding with RalGDS.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [10] MJ Smith, 2014, Integrated RAS signaling defined by parallel NMR detection of effectors and regulators, Nat Chem Biol source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    All G12X mutants show insensitivity to GAPs that accelerate hydrolysis.

    “All G12X mutants, however, show insensitivity to GAPs that accelerate hydrolysis [8].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [8] JC Hunter, 2015, Biochemical and structural analysis of common cancer-associated KRAS mutations, Mol Cancer Res source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    The G12D, G12R, and G12V mutants display highly reduced affinity to Raf, while the affinity of G12A is only moderately reduced.

    “The G12D, G12R, and G12V mutants display highly reduced affinity to Raf, while the affinity of G12A is only moderately reduced.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [8] JC Hunter, 2015, Biochemical and structural analysis of common cancer-associated KRAS mutations, Mol Cancer Res source

      Not checked in the free run.

  • Classified as probably not a substantial claim. May combine two claims.

    Premisecitation

    Switch-I displays highly dynamic characteristics manifested as two different states when GTP is bound to RAS, and the distribution between these states is altered in mutants.

    “Overall, switch-I displays highly dynamic characteristics manifested as two different states when GTP is bound to RAS, and the distribution between these states is altered in mutants [19–22].”
    Quote in the paperSubstantial claim0 of 4 sources checked
    • cites [19] M Geyer, 1996, Conformational transitions in p21ras and in its complexes with the effector protein Raf-RBD and the GTPase activating protein GAP, Biochemistry source

      Not checked in the free run.

    • cites [20] M Spoerner, 2001, Dynamic properties of the RAS switch I region and its importance for binding to effectors, Proc Natl Acad Sci USA source

      Not checked in the free run.

    • cites [21] M Spoerner, 2010, Conformational states of human rat sarcoma (RAS) protein complexed with its natural ligand GTP and their role for effector interaction and GTP hydrolysis, J Biol Chem source

      Not checked in the free run.

    • cites [22] M Spoerner, 2004, Perturbation of the conformational equilibria in RAS by selective mutations as studied by 31P NMR spectroscopy, FEBS Lett source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Every missense mutation at KRAS G12 (G12X) is oncogenic.

    “In fact, every missense mutation at G12 (G12X) is oncogenic.”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [3] JB Gibbs, 1984, Intrinsic GTPase activity distinguishes normal and oncogenic RAS p21 molecules, Proc. Natl. Acad. Sci. USA source

      Not checked in the free run.

    • cites [4] M Trahey, 1987, A cytoplasmic protein stimulates normal N-RAS p21 GTPase, but does not affect oncogenic mutants, Science source

      Not checked in the free run.

  • Classified as probably not a substantial claim.

    Interpretationreasoning

    If mutations took place randomly, G12D and G12S mutations should be the most abundant mutations as they are transition mutations.

    “If the mutations would take place randomly, G12D and G12S mutations should be the most abundant mutations as they are transition mutations (S2 Fig).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    Taken together, the results show for all mutants that the profile of their large-scale motions differs from wild-type regardless of the bound ligand, and that the profile is also unique to each mutant.

    “Taken together, the results show for all the mutants that the profile of their large-scale motions differs from wild-type regardless of the bound ligand, and that the profile is also unique to each mutant.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Studies of DNA-adduct formation by exogenous agents have resulted in adduct formation only in c.34G, and not in c.35G.

    “Interestingly, studies of DNA-adduct formation by exogenous agents have resulted in adduct formation only in c.34G, and not in c.35G [41,42].”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [41] Z Feng, 2002, Preferential DNA damage and poor repair determine RAS gene mutational hotspot in human cancer, J Natl Cancer Inst source

      Not checked in the free run.

    • cites [42] W Hu, 2003, Preferential carcinogen-DNA adduct formation at codons 12 and 14 in the human K-RAS gene and their possible mechanisms, Biochemistry source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    The oxidation of guanine by endogenous reactive oxygen species may result in DNA mutation.

    “The oxidation of guanine by endogenous reactive oxygen species may also result in DNA mutation [39].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [39] T Lindahl, 1993, Instability and decay of the primary structure of DNA, Nature source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Both guanines, the 5'G and the 3'G in a GGT-sequence, act as sites for frequent one-electron oxidation reactions and exhibit only a minor difference (0.05 eV) in their vertical ionization potential.

    “Both guanines, the 5’G and the 3’G in a GGT-sequence, are found to act as sites for frequent one-electron oxidation reactions, and they exhibit only a minor difference (0.05 eV) in their vertical ionization potential [37].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [37] A Bacolla, 2013, Guanine holes are prominent targets for mutation in cancer and inherited disease, PLoS Genet source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Oxidation can transform guanine to 7,8-dihydro-8-oxoguanine (8oxoG), which promotes especially the G>T transversion mutation.

    “The oxidation can further transform guanine to 7,8-dihydro-8-oxoguanine (8oxoG), which promotes especially the G>T transversion mutation [40]”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [40] SD Bruner, 2000, Structural basis for recognition and repair of the endogenous mutagen 8-oxoguanine in DNA, Nature source

      Not checked in the free run.

  • May combine two claims.

    Interpretationown data

    The conformation distribution of KRAS mutants includes conformations not occupied by wild-type, and these conformations are mutation specific.

    “This summarizes the main message: the conformation distribution of KRAS mutants includes conformations not occupied by wild-type, and these conformations are also mutation specific.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationcitation

    The 5'G position in the KRAS sequence (-TGGT-) represents the c.34G position, suggesting that the c.34G position is more susceptible for DNA-repair.

    “Correspondingly, this 5’G position in the KRAS sequence (-TGGT-) represents the c.34G position, thus suggesting that the c.34G position is more susceptible for DNA-repair.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Interpretationnone given

    There is no clear explanation why, on average, a mutation occurs five times more probably in c.35G than in c.34G.

    “For example, there is no clear explanation why, on average, a mutation occurs five times more probably in, e.g., c.35G than in c.34G.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S1 (asserted).

    Interpretationreasoning

    The biochemical and biophysical differences among mutants, resulting in signal-transducing differences, may explain, at least in part, the observed mutational bias.

    “Therefore, we hypothesize that the biochemical and biophysical differences among mutants, resulting in signal-transducing differences, may explain, at least in part, the observed mutational bias.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    The binding affinity of a specific KRAS mutant toward an effector protein can be increased or diminished, but the ability to bind still exists.

    “even though the binding affinity of a specific mutant toward an effector protein is increased or diminished, the ability to bind still exists [8].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [8] JC Hunter, 2015, Biochemical and structural analysis of common cancer-associated KRAS mutations, Mol Cancer Res source

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    The G12R mutation displays an advantage in the pancreas, while G12A is perhaps disadvantageous, given that there is a 9-fold preference for c.34G over c.35G in the mutation probability in the G>C mutations.

    “Similarly, the G12R mutation displays an advantage in the pancreas, while G12A is perhaps disadvantageous, given that there is a 9-fold preference for c.34G over c.35G in the mutation probability in the G>C mutations.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisenone given

    This is the first study to assess KRAS G12X mutation probabilities and to understand how they are associated with the observed mutation frequencies.

    “To the best of our knowledge, this is the first study to assess KRAS G12X mutation probabilities, and to understand how they are associated with the observed mutation frequencies.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

    Interpretationnone given

    One plausible explanation is that the 3D-environment in the DNA-sequence may aid the c.35G mutations to evade DNA-repair mechanisms.

    “One plausible explanation is that the 3D-environment in the DNA-sequence may aid the c.35G mutations to evade DNA-repair mechanisms.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Interpretationown data

    The PCA and interaction network analysis suggest that protein dynamics is altered among the systems, yet in some obscure manner.

    “As discussed above, the PCA and the interaction network analysis suggest that the protein dynamics is altered among the systems, yet in some obscure manner.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Interpretationreasoning

    The observed mutational effects on KRAS dynamics cannot be deduced to attenuate or amplify through HVR and cell membrane factors.

    “We therefore cannot deduce whether some of the observed mutational effects attenuate or amplify through these factors.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    It seems that either the advantage for G12D or the disadvantage for G12S, or both, exists in the pancreas, given that there is a 28-fold preference for G>A mutations for c.35G over c.34G.

    “For instance, it seems that either the advantage for G12D or the disadvantage for G12S, or both, exists in the pancreas, given that there is a 28-fold preference for G>A mutations for c.35G over c.34G.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    The HVR and cell membrane have a substantial influence on KRAS dynamics and signaling.

    “we recognize that these elements have a substantial influence on KRAS dynamics [53,54] and signaling [55].”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites [53] P Prakash, 2016, Membrane orientation dynamics of lipid-modified small GTPases, Small GTPases

      Not checked in the free run.

    • cites [54] DK Simanshu, 2017, RAS proteins and their regulators in human disease, Cell source

      Not checked in the free run.

    • cites [55] Y Zhou, 2017, Lipid-sorting specificity encoded in K-RAS membrane anchor regulates signal output, Cell source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premiseown data

    The switch-II conformation appears to be closed in the effector protein complexes.

    “In fact, the switch-II conformation appears to be closed in the effector protein complexes (S1A Fig).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    The basis for the specific frequencies of KRAS G12X mutations is poorly understood, and the effects of these specific mutations on a molecular scale are not clear.

    “Although frequently observed in cancer, not only is the basis for the specific frequencies of KRAS G12X mutations poorly understood [47], but also the effects of these specific mutations on a molecular scale are not clear.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [47] AG Stephen, 2014, Dragging RAS back in the ring, Cancer Cell source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    A mutation in V14 (V14I) is one of the responsible mutations for Noonan syndrome and may predispose tumor development.

    “a mutation in this position, V14I, is found to be one of the responsible mutations for the Noonan syndrome [57,58] and may also predispose tumor development [59].”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites [57] S Schubbert, 2006, Germline KRAS mutations cause Noonan syndrome, Nat Genet source

      Not checked in the free run.

    • cites [58] I Hernández-Porras, 2014, K-RasV14I recapitulates Noonan syndrome in mice, Proc Natl Acad Sci USA source

      Not checked in the free run.

    • cites [59] I Hernández-Porras, 2016, K-Ras(V14I) -induced Noonan syndrome predisposes to tumour development in mice, J Pathol source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Interpretationcitation

    The difference in mutant dynamics, such as G12V's inability to reach metastable states I and V, may reflect differences observed in RAS effector protein binding.

    “We consider that the difference in the mutant dynamics, for instance the G12V dynamics with its inability to reach the metastable states I and V (Fig 5), may reflect the differences observed in the RAS effector protein binding [8,10].”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [8] JC Hunter, 2015, Biochemical and structural analysis of common cancer-associated KRAS mutations, Mol Cancer Res source

      Not checked in the free run.

    • cites [10] MJ Smith, 2014, Integrated RAS signaling defined by parallel NMR detection of effectors and regulators, Nat Chem Biol source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductreasoning

    For PCA, the first 300 ns were discarded and only the last 1.7 μs of each simulated system were used to remove potential bias of the starting GDP crystal structure.

    “For PCA, we discarded the first 300 ns and used only the last 1.7 μs of each simulated system, to remove the potential bias of the starting GDP crystal structure from the results.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Interpretationcitation

    The altered dynamics may cause a conformational change of KRAS on the membrane, resulting in occluded conformation from a specific effector, alter dimer formation, or affect KRAS nanoclustering.

    “The altered dynamics may cause a conformational change of KRAS on the membrane, resulting in occluded conformation from a specific effector [23], alter the dimer formation [64], or affect KRAS nanoclustering [65].”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites [23] MT Mazhab-Jafari, 2015, Oncogenic and RASopathy-associated K-RAS mutations relieve membrane-dependent occlusion of the effector-binding site, Proc Natl Acad Sci USA source

      Not checked in the free run.

    • cites [64] X Nan, 2015, Ras-GTP dimers activate the mitogen-activated protein kinase (MAPK) pathway, Proc Natl Acad Sci USA source

      Not checked in the free run.

    • cites [65] R Nussinov, 2018, Is Nanoclustering essential for all oncogenic KRas pathways? Can it explain why wild-type KRas can inhibit its oncogenic variant?, Semin Cancer Biol

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Interpretationreasoning

    The absence of dynamic metastable states in crystal structures is due to the switch regions being disordered if there are no crystal contacts to the switches.

    “This is due to the fact that in the structures the switch regions are disordered, if there are no crystal contacts to the switches.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationnone given

    The mutation inflicts individual changes in the protein dynamics, affecting the allosteric communication network that conveys the shift in dynamics to remote sites within KRAS.

    “the mutation inflicts individual changes in the protein dynamics, affecting the allosteric communication network that conveys the shift in dynamics to the remote sites within KRAS.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Interpretationreasoning

    These metastable states may be important for specific effector protein binding and pathway activation.

    “These states may be important for specific effector protein binding and pathway activation.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductreasoning

    The first three N-terminal and last five C-terminal residues were excluded from PCA to reduce noise from flexible terminal regions.

    “To reduce noise from the flexible terminal regions, we excluded from the analysis the first three residues from the N-terminal and the last five residues from the C-terminal.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Premisecitation

    The KRAS G12X mutation data was collected from COSMIC database v.79.

    “The KRAS G12X mutation data was collected from COSMIC database v.79 (http://cancer.sanger.ac.uk/cosmic/) [2].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [2] SA Forbes, 2017, COSMIC: somatic cancer genetics at high-resolution, Nucleic Acids Res source

      Not checked in the free run.

  • Classified as probably not a substantial claim. May combine two claims.

    Study design and conductown data

    Each system was simulated for 2 μs with 5-10 independent replicas, giving individual system simulation time of 10-20 μs and total simulation time of 170 μs.

    “Each system was simulated for 2 μs with 5–10 independent replicas, such that the individual system simulation time was 10–20 μs and the total simulation time was 170 μs.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Interpretationcitation

    The observed metastable states may correspond to the first steps in the effector protein binding process.

    “the observed metastable states may correspond to the first steps in the effector protein binding process.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S4 (based).

    Premisenone given

    More complex mutations and their probabilities (e.g., adjacent double substitutions) are not predictable with existing knowledge.

    “This choice was made based on the fact that more complex mutations and their probabilities (e.g., adjacent double substitutions) are not predictable with the existing knowledge.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationreasoning

    A more stable intramolecular D154-R161 salt-bridge within the monomer could hinder dimer formation in GDP-bound KRAS, whereas a more unstable intramolecular salt-bridge in GTP-bound KRAS could promote intermolecular salt-bridges and dimer formation.

    “This suggests that the more stable intramolecular salt-bridge within the monomer residues could hinder the dimer formation in GDP-bound KRAS, whereas due to the change in the dynamics on the site in GTP-bound KRAS, the more unstable intramolecular salt-bridge could promote the formation of intermolecular salt-bridges among these residues, and thus dimer formation.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S2 (checkable), S3 (load-bearing).

    Interpretationnone given

    Multiple other aspects related to the altered dynamics may also affect KRAS mediated signaling.

    “multiple other aspects related to the altered dynamics may also affect KRAS mediated signaling.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Study design and conductreasoning

    Only single nucleotide substitutions were included in the assessment of KRAS G12X mutation probability.

    “In our assessment of the KRAS G12X mutation probability, we included only single nucleotide substitutions.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Interpretationown data

    The exact characteristics of interactions cannot be derived from the interaction frequency analysis, only their frequencies.

    “the exact characteristics of these interactions, unfortunately, cannot be derived from this analysis, only their frequencies.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Simple protein complexes assemble generally via a single pathway.

    “simple protein complexes assemble generally via a single pathway [62]”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [62] JA Marsh, 2015, Structure, dynamics, assembly, and evolution of protein complexes, Annu Rev Biochem source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Interpretationreasoning

    Even when a mutant such as G12D displays the same interaction frequencies as wild-type, the characteristics of the interactions may still differ.

    “even though a mutant, such as G12D, displays the same interaction frequencies as the wild-type, the characteristics of the interactions may still differ”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductcitation

    The number of clusters in k-means was set as √N, as recommended.

    “The number of clusters in k-means were set as √N, as recommended in [46].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [46] MK Scherer, 2015, PyEMMA 2: A software package for estimation, validation and analysis of Markov models, J Chem Theory Comp source

      Not checked in the free run.

95 of 95

  • The paper relies on this without support.

    Local DNA sequences have a major influence on the mutation probability.

    “Since local DNA-sequences have clearly a major influence on the mutation probability, a sequence-dependent basis for the observed non-random mutations may exist.”

  • The paper relies on this without support.

    The COSMIC database data are representative of KRAS G12X mutation frequencies in human cancers.

    “First, to perceive up-to-date data of KRAS G12X missense mutation frequencies, we compiled data from the Catalogue of Somatic Mutations in Cancer [2].”

  • The paper relies on this without support.

    The MD simulations and Markov state modeling accurately model the long-timescale conformational dynamics of KRAS.

    “To clarify the underlying causes for the nonrandom probabilities, we conducted extensive atomistic molecular dynamics simulations (170 μs) to study the differences of G12X mutations on a molecular level.”

  • The paper relies on this without support.

    KRAS G12X mutation has an effect on intrinsic GTPase activity and causes insensitivity for GAPs.

    “It has been clearly shown that the KRAS G12X mutation has an effect on the intrinsic GTPase activity and that it causes insensitivity for GAPs.”
4 of 4

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • Additional biochemical, structural, and signaling distinctions have been identified between different mutant KRAS alleles, including between those at the same amino acid position.

    “Additional biochemical, structural, and signaling distinctions have been identified between different mutant alleles, including between those at the same amino acid position4,8,13–20.”

    From The origins and genetic interactions of KRAS mutations are allele- and tissue-specific (2021) · cites this paper as [20]

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

Each contradiction between two passages listed here was confirmed by an independent check.

  • G12R group 1 occupancy conflicts with its component states

    Minor

    Group 1 consists of states I and V and is reported as 24% for G12R. But G12R occupies state I at 6%, while state V is reported as 12-16% for the other mutants, including G12R; together they can account for at most 22%.

  • State IV is described as both highly and sparsely populated

    Minor

    The paper includes state IV among the most populated states, but earlier explicitly places it among the less populated states I-IV.

2 of 2

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Related

Authors
Pantsar, T., Rissanen, S., Dauch, D., Laitinen, T., Vattulainen, I., & Poso, A.
Affiliations
  • University of Eastern Finland (FI)
  • Tampere University of Technology (FI)
  • Universitätsklinikum Tübingen (DE)
  • University of Helsinki (FI)
Corresponding author
  • Pantsar, Tatu
  • Poso, Antti
ORCID
Venue
PLOS Computational Biology
Publisher
Public Library of Science (PLoS)
Published
2018-09-10
Volume
14
Issue
9
Pages
e1006458
ISSN
  • 1553-734X (ISSN-L)
  • 1553-7358
DOI
10.1371/journal.pcbi.1006458
PMC
PMC6147662
OpenAlex
W2891963960
Semantic Scholar
3b130b91d2e0fe040dd11cd7d9e647e8ee0ad17d
Cited by
85
References
87
Citations by year
2026: 4 · 2025: 10 · 2024: 9 · 2023: 19 · 2022: 11 · 2021: 9 · 2020: 13 · 2019: 10
FWCI
2.84
Type
journal article
Language
en
Update notices
new_version (2018-09-20)
Open access
Licence
Locations
  • PLoS Computational Biology (publishedVersion, cc-by)
  • Europe PMC (PubMed Central) (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • RePEc: Research Papers in Economics (submittedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
  • UEF eRepo (University of Eastern Finland) (submittedVersion, cc-by)
  • Figshare (submittedVersion, cc-by)
  • Työväentutkimus Vuosikirja (submittedVersion)
  • Tampere University Institutional Repository (Tampere University) (submittedVersion, cc-by)
Funders
  • Academy of Finland (276509, 307415)
  • Biocenter Finland/DDCB
  • European Research Council (290974)
  • Tampereen Teknillinen Yliopisto (FI) (Graduate School Program)
  • Alfred Kordelinin Säätiö
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Kinase Regulation and GTPase Signaling
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Structure and Dynamics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Tyrosine Phosphatases
Keywords
KRAS, missense mutations, mutational bias, mutation frequency, oncogenic KRAS, allosteric communication, protein conformational dynamics, switch regions, effector binding, atomistic molecular dynamics
MeSH terms
DNA Mutational Analysis, Guanosine Diphosphate, Guanosine Triphosphate, Humans, Ligands, Markov Chains, Molecular Conformation, Mutation, Neoplasms, Probability, Genes, ras, Proto-Oncogene Proteins p21(ras), Mutation, Missense, Principal Component Analysis, Molecular Dynamics Simulation, Hydrophobic and Hydrophilic Interactions

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Claims read by deepseek-v4.1-flash under definition version 3 (extract-v3) on 2026-10-07.

Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-07.

Statistics and arithmetic are recomputed, not judged.

Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.

What counts as a claim ›How we measure confidence ›How we check this ›

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