CoChat
Sign Up
Cited by 48Antimicrobial Peptides and ActivitiesProtein Structure and DynamicsChemical Synthesis and Analysis

Structural and Functional Characterization of a Multifunctional Alanine-Rich Peptide Analogue from Pleuronectes americanus

Open access
Migliolo, Ludovico, Silva, Osmar N., Silva, Paula A., Costa, Maysa P., Costa, Carolina R., Nolasco, Diego O., Barbosa, João A. R. G., Silva, Maria R. R. +6 more·2012·PLoS ONE
DOI 10.1371/journal.pone.0047047PMC PMC3466273
Open this paper in CoChat — free accountOpen PDF

Abstract

Recently, defense peptides that are able to act against several targets have been characterized. The present work focuses on structural and functional evaluation of the peptide analogue Pa-MAP, previously isolated as an antifreeze peptide from Pleuronectes americanus. Pa-MAP showed activities against different targets such as tumoral cells in culture (CACO-2, MCF-7 and HCT-116), bacteria (Escherichia coli ATCC 8739 and Staphylococcus aureus ATCC 25923), viruses (HSV-1 and HSV-2) and fungi (Candida parapsilosis ATCC 22019, Trichophyton mentagrophytes (28d&E) and T. rubrum (327)). This peptide did not show toxicity against mammalian cells such as erythrocytes, Vero and RAW 264.7 cells. Molecular mechanism of action was related to hydrophobic residues, since only the terminal amino group is charged at pH 7 as confirmed by potentiometric titration. In order to shed some light on its structure-function relations, in vitro and in silico assays were carried out using circular dichroism and molecular dynamics. Furthermore, Pa-MAP showed partial unfolding of the peptide changes in a wide pH (3 to 11) and temperature (25 to 95°C) ranges, although it might not reach complete unfolding at 95°C, suggesting a high conformational stability. This peptide also showed a conformational transition with a partial α-helical fold in water and a full α-helical core in SDS and TFE environments. These results were corroborated by spectral data measured at 222 nm and by 50 ns dynamic simulation. In conclusion, data reported here show that Pa-MAP is a potential candidate for drug design against pathogenic microorganisms due to its structural stability and wide activity against a range of targets.

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

Integrity signals

Registry

Crossref · 10.1371/journal.pone.0047047

Cites

1 of 1 references verified0 citation errors · 0 not checkable

Statistics

Claims the paper makes

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

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?

  • Other AMPs such as Pa-MAP1, PGLa, and LL37 show similar structural transitions.

    “This idea was reinforced by the curiously parallel behavior observed here for other linear AMPs, including PGLa, a member of the magainin family [22], and LL37 [23]”

    From Structural insights into <i>Cn</i>‐AMP1, a short disulfide‐free multifunctional peptide from green coconut water (2015) · cites this paper as [21]

    The Discussion (page unavailable) describes a medium-dependent helix-to-coil transition for Pa-MAP, supporting that part of the claim. Our copy calls the peptide Pa-MAP, not Pa-MAP1, and does not establish comparable transitions for PGLa or LL37. (LLM, conf=0.86)

    Experimental — accuracy not yet measured

Self-check

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

The self-check needs the paper’s full text.

Cites

Flagged first, then by citations

Research works printed but not linked: 78

  1. [1]Nobeli I, Favia AD, Thornton JM (2009) Protein promiscuity and its implications for biotechnology. Nat Biotechnol 27: 157– 167. 19204698
  2. [2]Bechinger B (2010) Insights into themechanisms of action of host defence peptides from biophysical and structural investigations. Journal of Peptide Science 17: 306– 314. 21360627
  3. [3]Kuiper MJ, Fecondo JV, Wong MG (2002) Rational design of α-helical antifreeze peptides. J Pept Res 59: 1– 8. 11906602
  4. [4]Fletcher GL, Hew CL, Davies PL (2001) Antifreeze proteins of teleost fishes. Annu Rev Physiol 63: 359– 390. 11181960
  5. [5]Gong Z, Ewart KV, Hu Z, Fletcher GL, Hew CL (1996) Skin antifreeze protein genes of the winter flounder, Pleuronectes americanus, encode distinct and active polypeptides without the secretory signal and prosequences. J Biol Chem 271: 4106– 4112. 8626748
  6. [6]Hew CL, Joshi S, Wang NC, Kao MH, Ananthanarayanan VS (1985) Structures of shorthorn sculpin antifreeze polypeptides. Eur J Biochem 151: 167– 172. 4029130
  7. [7]Wang Z, Wang G (2004) APD: the Antimicrobial Peptide Database. Nucleic Acids Res 32: D590– 592. 14681488 PMC308759
  8. [8]Oren Z, Shai Y (1998) Mode of action of linear amphipathic α-helical antimicrobial peptides. Biopolymers 47: 451– 463. 10333737
  9. [9]Franco OL (2011) Peptide promiscuity: an evolutionary concept for plant defense. FEBS Lett 585: 995– 1000. 21402074
  10. [10]Hincha DK, DeVries AL, Schmitt JM (1993) Cryotoxicity of antifreeze proteins and glycoproteins to spinach thylakoid membranes–comparison with cryotoxic sugar acids. Biochim Biophys Acta 1146: 258– 264. 8452860
  11. [11]Tomczak MM, Hincha DK, Estrada SD, Wolkers WF, Crowe LM, et al (2002) A mechanism for stabilization of membranes at low temperatures by an antifreeze protein. Biophys J 82: 874– 881. 11806929 PMC1301896
  12. [12]Kun H, Minnes R, Mastai Y (2008) Effects antifreeze peptides on the thermotropic properties of a model membrane. J Bioenerg Biomembr 40: 389– 396. 18821004
  13. [13]Hancock RE, Sahl HG (2006) Antimicrobial and host-defense peptides as new anti-infective therapeutic strategies. Nat Biotechnol 24: 1551– 1557. 17160061
  14. [14]Holmberg N, Lilius G, Bulow L (1994) Artificial antifreeze proteins can improve NaCl tolerance when expressed in E. coli. FEBS Lett 349: 354– 358. 8050596
  15. [15]Sayers EW, Barrett T, Benson DA, Bolton E, Bryant SH, et al (2012) Database resources of the National Center for Biotechnology Information. Nucleic Acids Res 40: D13– 25. 22140104 PMC3245031
  16. [16]Hall T (1999) BioEdit: A user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symposium Series 4.
  17. [17]Thompson JD, Higgins DG, Gibson TJ (1994) CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 22: 4673– 4680. 7984417 PMC308517
  18. [21]Borenfreund E, Puerner JA (1985) Toxicity determined in vitro by morphological alterations and neutral red absorption. Toxicol Lett 24: 119– 124. 3983963
  19. [23]Markoulatos P, Georgopoulou A, Siafakas N, Plakokefalos E, Tzanakaki G, et al (2001) Laboratory diagnosis of common herpesvirus infections of the central nervous system by a multiplex PCR assay. J Clin Microbiol 39: 4426– 4432. 11724856 PMC88560
  20. [24]Reed LJ, Muench H (1938) A simple method of estimating fifty per cent endpoints. The American Journal of Hygiene 27: 5.
  21. [25]Simões C, Castro T, Cordeiro L, Albarello N, Mansur E, et al (2010) Antiviral activity of Cleome rosea extracts from field-grown plants and tissue culture-derived materials against acyclovir-resistant Herpes simplex viruses type 1 (ACVr-HSV-1) and type 2 (ACVr-HSV-2). World Journal Microbiology Biotechnology 26: 93– 99.
  22. [26]Nishimura T, Toku H, Fukuyasu H (1977) Antiviral compounds. XII. Antiviral activity of amidinohydrazones of alkoxyphenyl-substituted carbonyl compounds against influenza virus in eggs and in mice. Kitasato Arch Exp Med 50: 39– 46. 616473
  23. [27]Morrisett JD, David JS, Pownall HJ, Gotto AM Jr (1973) Interaction of an apolipoprotein (apoLP-alanine) with phosphatidylcholine. Biochemistry 12: 1290– 1299. 4348832
  24. [28]Eswar N, Webb B, Marti-Renom MA, Madhusudhan MS, Eramian D, et al.. (2006) Comparative protein structure modeling using Modeller. Curr Protoc Bioinformatics Chapter 5: Unit 5 6. PMC4186674 18428767
  25. [29]Wiederstein M, Sippl MJ (2007) ProSA-web: interactive web service for the recognition of errors in three-dimensional structures of proteins. Nucleic Acids Res 35: W407– 410. 17517781 PMC1933241
  26. [30]Sumathi K, Ananthalakshmi P, Roshan MN, Sekar K (2006) 3dSS: 3D structural superposition. Nucleic Acids Res 34: W128– 132. 16844975 PMC1538824
  27. [32]Dolinsky TJ, Nielsen JE, McCammon JA, Baker NA (2004) PDB2PQR: an automated pipeline for the setup of Poisson–Boltzmann electrostatics calculations. Nucleic Acids Research 32: W665– W667. 15215472 PMC441519
  28. [33]Wilkins MR, Gasteiger E, Bairoch A, Sanchez JC, Williams KL, et al (1999) Protein identification and analysis tools in the ExPASy server. Methods Mol Biol 112: 531– 552. 10027275
  29. [34]Lindahl E, Hess B, van der Spoel D (2001) GROMACS 3.0: a package for molecular simulation and trajectory analysis. Journal of Molecular Modeling 7: 306– 317.
  30. [36]Miyamoto S, Kollman P (1992) SETTLE: an analytical version of the SHAKE and RATTLE algorithms for rigid water models. Journal of Computational Chemistry 13: 952– 962.
  31. [37]Hess B, Bekker H, Berendsen H, Fraaije J (1997) LINCS: a linear constraint solver for molecular simulations. Journal of Computational Chemistry 18: 1463– 1472.
  32. [38]Liepinsh E, Otting G, Harding MM, Ward LG, Mackay JP, et al (2002) Solution structure of a hydrophobic analogue of the winter flounder antifreeze protein. Eur J Biochem 269: 1259– 1266. 11856360
  33. [39]Rost B, Yachdav G, Liu J (2004) The PredictProtein server. Nucleic Acids Res 32: W321– 326. 15215403 PMC441515
  34. [40]Geourjon C, Deleage G (1995) SOPMA: significant improvements in protein secondary structure prediction by consensus prediction from multiple alignments. Comput Appl Biosci 11: 681– 684. 8808585
  35. [41]Khameneh ZR, Sepehrvand N, Masudi S, Taghizade-Afshari A (2010) Seroprevalence of HTLV-1 among kidney graft recipients: a single-center study. Exp Clin Transplant 8: 146– 149. 20565372
  36. [42]Cutler DM (2007) The lifetime costs and benefits of medical technology. J Health Econ 26: 1081– 1100. 18023900
  37. [43]Foubister V (2003) New mode of intervention in sepsis treatment. Drug Discov Today 8: 610– 612. 12867140
  38. [44]Carrillo-Munoz AJ, Giusiano G, Ezkurra PA, Quindos G (2006) Antifungal agents: mode of action in yeast cells. Rev Esp Quimioter 19: 130– 139. 16964330
  39. [45]Hilpert K, Volkmer-Engert R, Walter T, Hancock RE (2005) High-throughput generation of small antibacterial peptides with improved activity. Nat Biotechnol 23: 1008– 1012. 16041366
  40. [46]Pelegrini PB, Franco OL (2005) Plant gamma-thionins: novel insights on the mechanism of action of a multi-functional class of defense proteins. Int J Biochem Cell Biol 37: 2239– 2253. 16084753
  41. [47]Pinto MF, Fensterseifer IC, Migliolo L, Sousa DA, de Capdville G, et al (2012) Identification and structural characterization of novel cyclotide with activity against an insect pest of sugar cane. J Biol Chem 287: 134– 147. 22074926 PMC3249065
  42. [48]Mandal SM, Dey S, Mandal M, Sarkar S, Maria-Neto S, et al (2009) Identification and structural insights of three novel antimicrobial peptides isolated from green coconut water. Peptides 30: 633– 637. 19111587
  43. [49]Silva ON, Porto WF, Migliolo L, Mandal SM, Garcês DG, et al. (2012) CnAMP-1: a new promiscuous peptide with potential for microbial infections treatment. Biopolymers Journal In press. 23193596
  44. [50]Zasloff M, Martin B, Chen HC (1988) Antimicrobial activity of synthetic magainin peptides and several analogues. Proc Natl Acad Sci U S A 85: 910– 913. 3277183 PMC279666
  45. [51]Yi N, Li N (2010) Transient expression of chicken antimicrobial peptides by mouse mammary carcinoma cells C127. Protein Pept Lett 17: 1517– 1523. 20937033
  46. [52]Baek JH, Ji Y, Shin JS, Lee S, Lee SH (2011) Venom peptides from solitary hunting wasps induce feeding disorder in lepidopteran larvae. Peptides 32: 568– 572. 21184791
  47. [53]Lee CC, Sun Y, Qian S, Huang HW (2011) Transmembrane pores formed by human antimicrobial peptide LL-37. Biophys J 100: 1688– 1696. 21463582 PMC3072607
  48. [54]Vad BS, Bertelsen K, Johansen CH, Pedersen JM, Skrydstrup T, et al (2010) Pardaxin permeabilizes vesicles more efficiently by pore formation than by disruption. Biophys J 98: 576– 585. 20159154 PMC2820650
  49. [55]Chang WT, Pan CY, Rajanbabu V, Cheng CW, Chen JY (2011) Tilapia (Oreochromis mossambicus) antimicrobial peptide, hepcidin 1–5, shows antitumor activity in cancer cells. Peptides 32: 342– 352. 21093514
  50. [56]Park NG, Silphaduang U, Moon HS, Seo JK, Corrales J, et al (2011) Structure-activity relationships of piscidin 4, a piscine antimicrobial peptide. Biochemistry 50: 3288– 3299. 21355570
  51. [57]Park CB, Lee JH, Park IY, Kim MS, Kim SC (1997) A novel antimicrobial peptide from the loach, Misgurnus anguillicaudatus. FEBS Lett 411: 173– 178. 9271200
  52. [58]Douglas SE, Gallant JW, Liebscher RS, Dacanay A, Tsoi SC (2003) Identification and expression analysis of hepcidin-like antimicrobial peptides in bony fish. Dev Comp Immunol 27: 589– 601. 12697315
  53. [59]Subramanian S, Ross NW, MacKinnon SL (2009) Myxinidin, a novel antimicrobial peptide from the epidermal mucus of hagfish, Myxine glutinosa L. Mar Biotechnol (NY). 11: 748– 757. 19330556
  54. [60]Broekman DC, Zenz A, Gudmundsdottir BK, Lohner K, Maier VH, et al (2011) Functional characterization of codCath, the mature cathelicidin antimicrobial peptide from Atlantic cod (Gadus morhua). Peptides 32: 2044– 2051. 21945422
  55. [61]Mandal SM, Migliolo L, Das S, Mandal M, Franco OL, et al (2012) Identification and characterization of a bactericidal and proapoptotic peptide from Cycas revoluta seeds with DNA binding properties. J Cell Biochem 113: 184– 193. 21882228
  56. [62]Mandal SM, Migliolo L, Franco OL, Ghosh AK (2011) Identification of an antifungal peptide from Trapa natans fruits with inhibitory effects on Candida tropicalis biofilm formation. Peptides 32: 1741– 1747. 21736910
  57. [63]Fehlbaum P, Bulet P, Chernysh S, Briand JP, Roussel JP, et al (1996) Structure-activity analysis of thanatin, a 21-residue inducible insect defense peptide with sequence homology to frog skin antimicrobial peptides. Proc Natl Acad Sci U S A 93: 1221– 1225. 8577744 PMC40060
  58. [64]Lee WH, Li Y, Lai R, Li S, Zhang Y, et al (2005) Variety of antimicrobial peptides in the Bombina maxima toad and evidence of their rapid diversification. Eur J Immunol 35: 1220– 1229. 15770703
  59. [65]Chang SL, Griesgraber G, Wagner CR (2001) Comparison of the antiviral activity of hydrophobic amino acid phosphoramidate monoesters of 2'3'-dideoxyadenosine (DDA) and 3'-azido-3'-deoxythymidine (AZT). Nucleosides Nucleotides Nucleic Acids 20: 1571– 1582. 11554546
  60. [66]Lee DG, Park Y, Jin I, Hahm KS, Lee HH, et al (2004) Structure-antiviral activity relationships of cecropin A-magainin 2 hybrid peptide and its analogues. J Pept Sci 10: 298– 303. 15160842
  61. [67]Pan CY, Chen JY, Cheng YS, Chen CY, Ni IH, et al (2007) Gene expression and localization of the epinecidin-1 antimicrobial peptide in the grouper (Epinephelus coioides), and its role in protecting fish against pathogenic infection. DNA Cell Biol 26: 403– 413. 17570764
  62. [68]Zhang L, Falla T, Wu M, Fidai S, Burian J, et al (1998) Determinants of recombinant production of antimicrobial cationic peptides and creation of peptide variants in bacteria. Biochem Biophys Res Commun 247: 674– 680. 9647752
  63. [69]Huang YB, Wang XF, Wang HY, Liu Y, Chen Y (2012) Studies on mechanism of action of anticancer peptides by modulation of hydrophobicity within a defined structural framework. Mol Cancer Ther 10: 416– 426. 21252288
  64. [70]Wang G, Li X, Wang Z (2009) APD2: the updated antimicrobial peptide database and its application in peptide design. Nucleic Acids Res 37: D933– 937. 18957441 PMC2686604
  65. [71]Rozek T, Wegener KL, Bowie JH, Olver IN, Carver JA, et al (2000) The antibiotic and anticancer active aurein peptides from the Australian Bell Frogs Litoria aurea and Litoria raniformis the solution structure of aurein 1.2. Eur J Biochem 267: 5330– 5341. 10951191
  66. [72]Chernysh S, Kim SI, Bekker G, Pleskach VA, Filatova NA, et al (2002) Antiviral and antitumor peptides from insects. Proc Natl Acad Sci U S A 99: 12628– 12632. 12235362 PMC130511
  67. [73]Zeng XC, Wang S, Nie Y, Zhang L, Luo X (2011) Characterization of BmKbpp, a multifunctional peptide from the Chinese scorpion Mesobuthus martensii Karsch: Gaining insight into a new mechanism for the functional diversification of scorpion venom peptides. Peptides 33: 44– 51. 22115565
  68. [74]Verkleij AJ, Zwaal RF, Roelofsen B, Comfurius P, Kastelijn D, et al (1973) The asymmetric distribution of phospholipids in the human red cell membrane. A combined study using phospholipases and freeze-etch electron microscopy. Biochim Biophys Acta 323: 178– 193. 4356540
  69. [75]Turner JD, Rouser G (1970) Precise quantitative determination of human blood lipids by thin-layer and triethylaminoethylcellulose column chromatography. I. Erythrocyte lipids. Anal Biochem 38: 423– 436. 5493067
  70. [76]Kreil G (1973) Biosynthesis of melittin, a toxic peptide from bee venom. Amino-acid sequence of the precursor. Eur J Biochem 33: 558– 566. 4571501
  71. [77]Werkmeister JA, Kirkpatrick A, McKenzie JA, Rivett DE (1993) The effect of sequence variations and structure on the cytolytic activity of melittin peptides. Biochim Biophys Acta 1157: 50– 54. 8499479
  72. [78]Domadia PN, Bhunia A, Ramamoorthy A, Bhattacharjya S (2011) Structure, interactions, and antibacterial activities of MSI-594 derived mutant peptide MSI-594F5A in lipopolysaccharide micelles: role of the helical hairpin conformation in outer-membrane permeabilization. J Am Chem Soc 132: 18417– 18428. 21128620
  73. [79]Sung WS, Park SH, Lee DG (2008) Antimicrobial effect and membrane-active mechanism of Urechistachykinins, neuropeptides derived from Urechis unicinctus. FEBS Lett 582: 2463– 2466. 18570895
  74. [80]Jasanoff A, Fersht AR (1994) Quantitative determination of helical propensities from trifluoroethanol titration curves. Biochemistry 33: 2129– 2135. 8117669
  75. [82]Ding T, Li R, Zeitler JA, Huber TL, Gladden LF, et al (2010) Terahertz and far infrared spectroscopy of alanine-rich peptides having variable ellipticity. Opt Express 18: 27431– 27444. 21197019
  76. [83]Roccatano D, Colombo G, Fioroni M, Mark AE (2002) Mechanism by which 2,2,2-trifluoroethanol/water mixtures stabilize secondary-structure formation in peptides: a molecular dynamics study. Proc Natl Acad Sci U S A 99: 12179– 12184. 12196631 PMC129418
  77. [84]Jalili S, Akhavan M (2010) Molecular dynamics simulation study of association in trifluoroethanol/water mixtures. J Comput Chem 31: 286– 294. 19475633
  78. [85]Li H, Monien BH, Fradinger EA, Urbanc B, Bitan G (2010) Biophysical characterization of Aβ42 C-terminal fragments: inhibitors of Aβ42 neurotoxicity. Biochemistry 49: 1259– 1267. 20050679 PMC2831638

Cited by

Sorted by citations

Authors
Migliolo, L., Silva, O. N., Silva, P. A., Costa, M. P., Costa, C. R., Nolasco, D. O., Barbosa, J. A. R. G., Silva, M. R. R., Bemquerer, M. P., Lima, L. M. P., Romanos, M. T. V., Freitas, S. M., Magalhães, B. S., & Franco, O. L.
Affiliations
  • Universidade Católica de Brasília (BR)
  • Universidade Federal de Juiz de Fora (BR)
  • Universidade Federal do Rio de Janeiro (BR)
  • Universidade Federal de Goiás (BR)
  • Universidade de Brasília (BR)
  • Embrapa (BR)
ORCID
Venue
PLoS ONE
Publisher
Public Library of Science (PLoS)
Published
2012-10-08
Volume
7
Issue
10
Pages
e47047
ISSN
1932-6203 (ISSN-L)
DOI
10.1371/journal.pone.0047047
PMC
PMC3466273
OpenAlex
W2079161080
Cited by
48
References
1
Citations by year
2026: 3 · 2025: 6 · 2024: 2 · 2023: 2 · 2022: 5 · 2021: 4 · 2020: 5 · 2019: 1 · 2018: 3 · 2017: 3 · 2016: 7 · 2015: 1 · 2014: 3 · 2013: 3
FWCI
1.94
Type
journal article
Language
en
Open access
Licence
Locations
  • PLoS ONE (publishedVersion, cc-by)
  • Europe PMC (PubMed Central) (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • Americanae (AECID Library) (submittedVersion, other-oa)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
  • LA Referencia (Red Federada de Repositorios Institucionales de Publicaciones Científicas) (submittedVersion, other-oa)
Funders
  • Coordenação de Aperfeiçoamento de Pessoal de Nível Superior
  • Conselho Nacional de Desenvolvimento Científico e Tecnológico
  • Fundação de Apoio à Pesquisa do Distrito Federal
Topics
  • Life Sciences › Immunology and Microbiology › Microbiology › Antimicrobial Peptides and Activities
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Structure and Dynamics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Chemical Synthesis and Analysis
Keywords
winter flounder, antifreeze peptides, multifunctional peptides, antimicrobial activity, antiviral activity, antifungal activity, anticancer activity, structure-function relationship, conformational stability, alpha-helix, hydrophobic residues, molecular dynamics simulation, circular dichroism, drug design
MeSH terms
Alanine, Animals, Candida, Cell Line, Erythrocytes, Flounder, Humans, Peptides, Staphylococcus aureus, Trichophyton, Caco-2 Cells, HCT116 Cells

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.

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 ›

Are you an author, or do you see a problem with this page? Request a review