CoChat
Sign Up
Cited by 33Virus-based gene therapy researchRetinal Development and DisordersRNA Interference and Gene Delivery

Structure comparison of the chimeric AAV2.7m8 vector with parental AAV2

Open access
Bennett, Antonette, Keravala, Annahita, Makal, Victoria, Kurian, Justin, Belbellaa, Brahim, Aeran, Rangoli, Tseng, Yu-Shan, Sousa, Duncan +3 more·2020·Journal of Structural Biology
DOI 10.1016/j.jsb.2019.107433PMC PMC7058502
Open this paper in CoChat — free account

Abstract

The AAV2.7m8 vector is an engineered capsid with a 10-amino acid insertion in adeno-associated virus (AAV) surface variable region VIII (VR-VIII) resulting in the alteration of an antigenic region of AAV2 and the ability to efficiently transduce retina cells following intravitreal administration.

Integrity signals

Registry

Crossref · 10.1016/j.jsb.2019.107433

Cites

102 of 102 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

21 of 35 cited sources checked · 7 supported · 1 flagged · accuracy not yet measured

  • Premisecitation

    VR-VIII is part of an AAV2 antigenic epitope.

    “VR-VIII is part of an AAV2 antigenic epitope, and the difference is consistent with rAAV2.7m8’s escape from a known AAV2 monoclonal antibody, C37-B.”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites Gurda et al., 2013 source
  • Premisecitation

    The reported AAV2 HS binding residues are R484, R487, K532, R585, and R588, corresponding to R484, R487, K532, R585, and R598 in rAAV2.7m8.

    “The reported AAV2 HS binding residues are R484, R487, K532, R585, and R588 (Fig. 6C) (Kern et al., 2003; Opie et al., 2003) corresponding to residues R484, R487, K532, R585, and R598 in rAAV2.7m8.”
    Quote in the paperSubstantial claim1 of 2 sources checked
    • cites Kern et al., 2003 source
    • cites Opie et al., 2003 source
  • Interpretationcitation

    The lack of ordered density for VP1u, VP1/2 common region, and VP3 N-terminus is consistent with previous AAV2 and other AAV capsid structures.

    “The lack of VP1u (residues 1-137), VP1/2 (residues 138-202), and VP3 N-terminus (residues 203-216/218) ordering is consistent with the previous report for the AAV2 crystal structure as well as reports for other AAV capsid structures (reviewed in (Mietzsch et al., 2019)).”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites Mietzsch et al., 2019 source
  • Interpretationcitation

    The result agrees with the peptide insert sitting above the C37-B epitope, which was mapped to VR-VIII by cryo-EM of the AAV2-C37-B complex.

    “This observation is consistent with the positioning of the peptide insert above at the epitope mapped for C37-B to VR-VIII using cryo-EM of the AAV2-C37-B complex (Fig. 8B) (Gurda et al., 2013).”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites Gurda et al., 2013 source
      Experimental — accuracy not yet measured
  • Findingcitation

    In rAAV2.7m8, Y714 (Y704 in AAV2) and E563, two pH quartet residues, adopt the reported AAV8 pH4 conformation.

    “Two of the residues in the pH quartet, Y714 (Y704 in AAV2) and E563, are in the reported AAV8 pH4 conformation (Nam et al., 2011) in rAAV2.7m8, implying that rAAV2.7m8 is already primed for endosomal escape or that this capsid region samples at least two conformations.”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites Nam et al., 2011 source
  • May combine two claims. 32
  • Classified as probably not a substantial claim. 9
  • The quote only partly states this claim. 2

  • May combine two claims.

    Premisecitation

    At pH 7.4, the pH quartet residues of AAV2 adopt the same conformation as in AAV8, although E563 was less ordered.

    “In AAV2, all the pH quartet residues have the same conformational as in AAV8 at pH 7.4 (Nam et al., 2011), however, acidic residue E563 was less ordered.”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites Nam et al., 2011 source

      The Results and Discussion (Fig. 3; page not available in our copy) say that AAV2 Y704 has the same conformation as the equivalent AAV8 residue, but the equivalent arginine and glutamate residues are too far apart in AAV2 to make the interaction seen in AAV8. This contradicts the claim that all quartet residues have the same conformation. The passage does not say that E563 was less ordered. (LLM, conf=0.85)

      Checked by a reasoning model — may be wrong.

      Experimental — accuracy not yet measured

  • May combine two claims.

    Premisecitation

    rAAV2.7m8 crosses the inner limiting membrane in the retina, whereas other naturally occurring AAV serotypes including AAV2 penetrate the retina poorly.

    “Furthermore, rAAV2.7m8 was shown to cross the inner limiting membrane in the retina, where other naturally occurring AAV serotypes, including its parental AAV2, have poor retina penetration (Aartsen et al., 2010; Dalkara et al., 2009; Dalkara et al., 2013; Giove et al., 2010; Hellstrom et al., 2009).”
    Quote in the paperSubstantial claim1 of 3 sources checked
    • cites Aartsen et al., 2010 source

      The Discussion (page not provided) says the inner limiting membrane is a barrier to AAV2/6 penetration and notes a similar barrier for other AAV serotypes. It does not report that rAAV2.7m8 crosses the membrane or specifically establish poor retinal penetration by its parental AAV2. (LLM, conf=0.87)

      Experimental — accuracy not yet measured
    • cites Dalkara et al., 2009 source
    • cites Dalkara et al., 2013 source

  • May combine two claims.

    Premisecitation

    rAAV2.7m8 is an engineered AAV capsid with a ten-amino-acid insertion in VR-VIII, derived from AAV2 and isolated by directed evolution for increased mouse retina transduction after intravitreal administration.

    “This includes the recombinant AAV vector rAAV2.7m8 (Dalkara et al., 2013), which is derived from the natural serotype AAV2 and was isolated by directed evolution when screening for increased transduction of the mouse retina following intravitreal (IVT) administration.”
    • Also stated in: § INTRODUCTION · plain
      “In this vector, peptide LALGETTRPA is inserted after amino position R588 of AAV2, located in a capsid surface loop designated variable region VIII (VR-VIII) based on 3D structure comparison (Govindasamy et al., 2006).”
    Quote in the paperSubstantial claim1 of 2 sources checked
    • cites Dalkara et al., 2013 source
    • cites Govindasamy et al., 2006 source

      The Abstract describes AAV4 and AAV2 capsid structures, and the Discussion describes capsid regions that may tolerate insertions. Neither addresses rAAV2.7m8, a ten-amino-acid insertion, or directed evolution for mouse-retina transduction after intravitreal administration. Searches for rAAV2.7m8, directed evolution, and the VR-VIII insertion found no supporting passage in our copy. (LLM, conf=0.76)

      Experimental — accuracy not yet measured

  • Classified as probably not a substantial claim.

    Premisecitation

    Prior work characterized R389 as a member of the four-residue pH quartet (R389, H526, E563, Y704) that undergoes pH-induced conformational change, which is essential for viral infectivity.

    “R389 has a dual conformation in rAAV2.7m8 but not AAV2, and has been previously characterized as one of four residues (R389, H526, E563 and Y704 - pH quartet) that undergo pH-induced conformational change, an essential step in viral infectivity (Bartlett et al., 2000; Douar et al., 2001; Hansen et al., 2001; Nam et al., 2011; Nam et al., 2007).”
    Quote in the paperSubstantial claim2 of 3 sources checked
    • cites Bartlett et al., 2000 source

      The Results state that AAV requires passage through an acidic compartment for productive infection, and the Discussion reports that raising endosomal pH inhibits infection. The paper does not characterize R389, identify a four-residue pH quartet, or show that those residues undergo a pH-induced conformational change. (LLM, conf=0.87)

      Experimental — accuracy not yet measured
    • cites Douar et al., 2001 source
    • cites Hansen et al., 2001 source

      The Discussion (page unavailable) describes a low-pH-dependent AAV capsid modification that may help the virus reach the nucleus, but says its exact nature is undefined. The paper does not identify R389 or the four-residue pH quartet, and it says low pH alone is insufficient to cause the AAV modification. (LLM, conf=0.86)

      Experimental — accuracy not yet measured

  • May combine two claims.

    Interpretationcitation

    The rAAV2.7m8 insertion modifies the C37-B antibody binding site, creating an escape variant.

    “The rAAV2.7m8 insertion modified the C37-B antibody binding site (Gurda et al., 2013) creating an escape variant.”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites Gurda et al., 2013 source

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

    Premisecitation

    For AAV2, two critical residues required for binding HS, R585 and R588, reside within VR-VIII, and the 511-NGR-513 motif proposed to play a role in α5β1 co-receptor binding is flanked by VR-I and VR-IV.

    “For example, for AAV2, two critical residues required for binding HS, R585 and R588, reside within VR-VIII, and the 511-NGR-513 proposed to play a role in α5β1 co-receptor binding is flanked by VR-I and VR-IV (Asokan et al., 2006; Kern et al., 2003; Levy et al., 2009; Ng et al., 2010; Opie et al., 2003; Shen et al., 2015; Summerford and Samulski, 1998).”
    Quote in the paperNot a substantial claim2 of 3 sources checked
    • cites Asokan et al., 2006 source

      The Results identify the AAV2 NGR motif at positions 511–513 as a proposed integrin-binding sequence. The Discussion places it adjacent to R585 and R588, residues previously implicated in heparin binding. Our copy does not identify R585/R588 as lying within VR-VIII or describe the NGR motif as flanked by VR-I and VR-IV. (LLM, conf=0.86)

      Experimental — accuracy not yet measured
    • cites Kern et al., 2003 source
    • cites Levy et al., 2009 source

      The Introduction identifies residues 511-NGR-513 as an α5β1-binding motif, and Results and Discussion calls R585 and R588 the residues most critical for AAV2 heparin binding. The cited paper does not place R585 and R588 within VR-VIII or say that VR-I and VR-IV flank the NGR motif (no page numbers are available in our copy). (LLM, conf=0.87)

      Experimental — accuracy not yet measured

  • May combine two claims.

    Premisecitation

    A20's epitope runs from the 2/5-fold wall toward the 5-fold axes and does not share residues with the peptide.

    “While the A20 epitope is positioned from the 2/5-fold wall towards the 5-fold axes and shares no residues with the peptide (McCraw et al., 2012), the C37-B epitope is significantly overlapped by the position of the inserted peptide (Fig. 8B).”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites McCraw et al., 2012 source

      The Results and Discussion sections on the binding site and epitope place A20’s footprint across the plateau, wall, and canyon, supporting its general location. They do not establish that it extends toward the fivefold axes or compare its residues with the specific inserted peptide in the citing sentence; the paper discusses other peptide-insertion sites instead. Page numbers were not available in our copy. (LLM, conf=0.82)

      Experimental — accuracy not yet measured

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

    Premisecitation

    The pore at the 5-fold axes is postulated to be the site for externalization of the PLA2 enzyme within VP1u required for endosomal escape during infection and for genome packaging and release during infection.

    “The pore at the 5-fold axes is postulated to be the site for externalization of the PLA2 enzyme within VP1u required for endosomal escape during infection and for genome packaging and release during infection (Bleker et al., 2005; Grieger et al., 2007; Kronenberg et al., 2005; Sonntag et al., 2006).”
    Quote in the paperNot a substantial claim2 of 3 sources checked
    • cites Bleker et al., 2005 source

      The Abstract and Discussion (page numbers unavailable) propose that AAV2 fivefold pores expose the VP1 PLA2 domain during infection and may serve as channels for genome packaging. Our copy does not identify endosomal escape as the function of that exposure or propose that AAV2 genomes are released through those pores during infection; the Discussion describes pore-mediated genome release in other viruses. (LLM, conf=0.86)

      Experimental — accuracy not yet measured
    • cites Grieger et al., 2007 source
    • cites Kronenberg et al., 2005 source

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

    Premisecitation

    The 3-fold region, including the protrusions, contains receptor binding, infectivity, and antigenicity controlling residues.

    “The 3-fold region, including the protrusions, contain receptor binding, infectivity, and antigenicity controlling residues (reviewed in (Mietzsch et al., 2019)).”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites Mietzsch et al., 2019 source

      Section 2.2 describes variable loops at the 3-fold region as mediating interactions with receptors and antibodies; Sections 2.5 and 2.6 also identify receptor-binding sites and antibody epitopes on or around the protrusions. These passages support the receptor-binding and antigenicity parts, but the copy does not establish that residues in this region control infectivity. (LLM, conf=0.83)

      Experimental — accuracy not yet measured

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

    Premisecitation

    Among AAV serotypes, AAV2 has been studied most extensively, and every charged residue in its VP has been functionally characterized.

    “AAV2 is the most extensively studied AAV serotype, including the functional characterization of all charged residues in the VP (Choi et al., 2005; Grieger et al., 2006; Lochrie et al., 2006; Mietzsch et al., 2019; Wang et al., 2019; Wu et al., 2000).”
    Quote in the paperNot a substantial claim2 of 3 sources checked
    • cites Choi et al., 2005 source

      Section 1.2 calls AAV2 the best characterized and extensively studied naturally occurring serotype. Section 3.1 reports functional effects of mutations at specific charged capsid positions, but does not say that every charged residue in its VP proteins has been functionally characterized. The claim’s ‘all charged residues’ qualifier is therefore unsupported by this paper. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites Grieger et al., 2006 source
    • cites Lochrie et al., 2006 source

      The Abstract describes extensive prior work on AAV2 capsid mutations, and the Results report functional assays of mutants at 64 external-surface positions. But the Results say those positions cover only about 55% of the external surface; the paper does not establish that every charged residue in the VP proteins was functionally characterized, or explicitly compare how extensively all serotypes have been studied. The indexed passages provide section labels but no page numbers. (LLM, conf=0.85)

      Experimental — accuracy not yet measured

  • May combine two claims.

    Findingown data

    The structures of rAAV2.7m8 and AAV2 were determined by cryo-EM to 2.91 Å and 3.02 Å resolution, respectively.

    “The structures of rAAV2.7m8 and AAV2 were determined to 2.91 and 3.02 Å resolution, respectively.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    The rAAV2.7m8 and AAV2 structures are similar except at some side-chain orientations and at the VR-VIII/3-fold protrusion region containing the ten-amino-acid insertion.

    “A comparison to parental AAV2 (ordered from residues 217-735) showed the structures to be similar, except at some side-chains that had different orientations and, in VR-VIII containing the 10 amino acid insertion.”
    • Also stated in: § RESULTS AND DISCUSSION · plain
      “The rAAV2.7m8 and AAV2 structures are similar with the exception of the 3-fold protrusions containing the 7m8 insertion (Fig. 2).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    These basic residues are located in the 2-fold symmetry-related interface, conserved secondary structures, and facing the luminal surface of the capsid.

    “These basic residues are located in the 2-fold symmetry related interface, the conserved secondary structures, and facing the luminal surface of the capsid (Table 2).”
    Quote in the paperNot a substantial claim

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

    Findingown data

    The rAAV2.7m8 VP3 region was ordered from residue 219 to 745, while AAV2 was ordered from residue 217 to 735.

    “The rAAV2.7m8 was ordered from residue 219 to 745 (last C-terminal residue, VP1 numbering) while AAV2 was ordered from residue 217 to 735 (last C-terminal residue).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    At ≥3.0σ, side-chain density was observable up to residue N587 on the ascending arm and after R595 on the descending arm of the inserted loop.

    “At ≥3.0σ, side-chain density was observable for up to residue N587 on the ascending arm and after R595 on the descending arm of the inserted loop (Fig. 4A).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    A surface view of rAAV2.7m8 and AAV2 structures indicates that the insertion obscures access to some HS binding residues, although not R585 or R598.

    “However, a surface view of rAAV2.7m8 and AAV2 structures indicates that the insertion in the latter obscures access to some of the residues, although not R585 or R598 (Fig. 6C and D).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    The rAAV2.7m8 peptide insertion does not alter the position or structure of the HS binding residues R585 and R598, nor that of the other HS binding residues.

    “The rAAV2.7m8 peptide insertion does not alter the position or structure of the R585 and R598 (Fig. 6A and B) nor that of the other HS binding residues (not shown).”
    Quote in the paperSubstantial claim

  • May combine two claims. The quote only partly states this claim.

    Findingown data

    The overall VP topology and main-chain/secondary structure of rAAV2.7m8 are conserved with AAV2, except for a 10-residue insertion at the 3-fold protrusion.

    “The main-chain and secondary structure elements of rAAV2.7m8 and AAV2 are structurally conserved (Fig. 5A and 5B), except for the 10 residue insertion in AAV2.7m8 as described above.”
    • Also stated in: § SUMMARY · plain
      “In this case, the new vector’s structure retained the overall VP topology and capsid morphology of the parental AAV2 except at the 3-fold protrusion important for receptor binding and antigenicity.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    The wall of the 2-fold depression is formed by VR-VI, the 2/5-fold wall by VR-I, VR-III, and VR-IX, the top of the 5-fold channel by VR-II, and the protrusions surrounding the 3-fold axis by VR-IV, VR-V, and VR-VIII.

    “The wall of the 2-fold depression is formed by VR-VI, the 2/5-fold wall is formed by VR-I, VR-III, and VR-IX, the top of the 5-fold channel is formed by VR-II, and the protrusions surrounding the 3-fold axis is composed of VR-IV, VR-V, and VR-VIII (Fig. 2, 5B, and C).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    In rAAV2.7m8, C361 adopts an alternate conformation relative to AAV2 and interacts with H288 via extra density between them.

    “Residue C361 (Table 2) has an alternate conformation in rAAV2.7m8 compared to AAV2, and is involved in an interaction with H288 through additional density present between the two residues.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    The rAAV2.7m8 capsid maintains the parvovirus capsid morphology, including a depression at the icosahedral 2-fold axes and surrounding the 5-fold axes, protrusions surrounding the 3-fold axes, and a channel at the 5-fold axes.

    “The rAAV2.7m8 capsid maintains the capsid morphology of the parvoviruses that includes a depression at the icosahedral 2-fold axes and surrounding the 5-folds axes, protrusions surround the 3-fold axes, and a channel at the 5-fold axes (Fig. 2A).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    In rAAV2.7m8, density for residues T581-Q599 was evident at 1σ, but the three residues at the top of the loop, L590-E592, were not ordered at 2σ.

    “While the density for residues T581-Q599 was evident at 1σ in rAAV2.7m8, the three residues at the top of this loop, L590-E592, were not ordered at 2σ (Fig. 3D and 4).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Electron density was observed for the entire loop formed by the 7m8 insertion peptide, but side-chain density was resolved for only six of the ten amino acids, with the remaining residues observed only at a sigma threshold of 1.

    “Electron density was observed for the entire loop formed by the 7m8 insertion peptide; however, the side-chain density could only be resolved for six out the ten amino acids and the remaining residues were only observed at a sigma threshold of 1.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    The 10-amino-acid insertion in rAAV2.7m8 is located at the top of VR-VIII, which forms the side of the 3-fold protrusion facing the icosahedral 3-fold axis and extends further out than VR-IV, unlike in parental AAV2.

    “The 10 amino acid peptide insertion in rAAV2.7m8 is located at the top of VR-VIII (Fig. 5A and B) that forms the side of the 3-fold protrusion facing the icosahedral 3-fold axis that now extends further out than VR-IV, unlike in the parental AAV2 (Fig. 5A, B, 6A, and B).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    The rAAV2.7m8 VP structure conserved the secondary structure elements reported for parvoviruses, including an eight-stranded anti-parallel β-barrel core with the BIDG sheet facing the virus interior and the αA helix forming the wall of the 2-fold axes.

    “The rAAV2.7m8 VP structure conserved the secondary structure elements reported for parvoviruses. This includes an eight-stranded anti-parallel β-barrel core, with the BIDG sheet facing the virus interior, and the αA helix forming the wall of the 2-fold axes (Fig. 5A).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Critical residues R585 and R588/598 are positioned adjacent to each other from the ascending and descending arms of VR-VIII, and together with other residues at the base of the 3-fold protrusions form a basic patch on the capsid surface.

    “Critical residues R585 and R588/598, are positioned adjacent to each other from the ascending and descending arms, respectively, of VR-VIII, and together with the other residues, located at the base of the 3-fold protrusions, form a basic patch on the capsid surface (Fig. 6).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    The 7m8 insertion was predicted and shown to disrupt AAV2 binding to heparan sulfate proteoglycan.

    “This insertion was predicted and shown to disrupt the binding of AAV2 to its primary receptor, heparan sulfate proteoglycan (HSPG) (Dalkara et al., 2013).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Dalkara et al., 2013 source

  • May combine two claims.

    Interpretationreasoning

    For AAV2.7m8, the 3-fold protrusion is important for cellular trafficking because the vector can reach the retina after intravitreal delivery, unlike parental AAV2.

    “For AAV2.7m8, this region is also important for cellular trafficking, given its ability to reach the retina following intravitreal administration, a capability not shared by parental AAV2.”
    Quote in the paperSubstantial claim
    • Rests on an assumption: AAV2.7m8 can reach the retina after intravitreal administration, whereas parental AAV2 cannot.

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

    Study design and conductnone given

    The gold standard protocol was followed throughout the structure determination process.

    “The gold standard protocol was followed throughout the structure determination process.”
    Quote in the paperNot a substantial claim

  • The quote only partly states this claim.

    Interpretationown data

    The structural similarity of HS binding residues is consistent with rAAV2.7m8 retaining the ability to bind HS.

    “consistent with the ability of rAAV2.7m8 to still bind HS.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    The observation supports using peptide insertion both to change tissue tropism and to reduce antigenic reactivity.

    “This observation supports a two-pronged approach involving peptide insertion for altering tissue tropism as well as ablating antigenic reactivity.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    Residues with altered or dual conformations, despite the two viruses' sequence identity, lay in functional regions and in areas with conformational freedom.

    “Residues exhibiting altered or dual conformations, despite identity between the two viruses, were in functional regions and localized to areas with conformational freedom.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Study design and conductown data

    The purity of rAAV2.7m8 and AAV2 was confirmed by gel electrophoresis, and capsid integrity was confirmed by EM.

    “The purity of rAAV2.7m8 and AAV2 were confirmed by gel electrophoresis, and the integrity of the capsids was confirmed by EM (Fig. 1A).”
    Quote in the paperSubstantial claim

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

    Premisecitation

    Previous mutagenesis and biophysical studies reported that some of these residues are important in virus assembly, production, infectivity, and pH-induced changes during trafficking.

    “Previous mutagenesis and biophysical studies have reported the importance of some of these residues in virus assembly, production (Grieger et al., 2006; Wu et al., 2000), infectivity (Zhong et al., 2008), and pH-induced changes during trafficking (Nam et al., 2011; Salganik et al., 2012).”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites Grieger et al., 2006 source
    • cites Wu et al., 2000

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

    • cites Zhong et al., 2008 source

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

    Interpretationreasoning

    The additional R238 interaction possibly results in a more stable capsid compared with AAV2.

    “R238 of rAAV2.7m8 is involved in one additional interaction at the 2-fold axes possibly resulting in a more stable capsid compared to AAV2 (not shown).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisenone given

    The residues forming the nucleotide binding pocket are V418, P419, H639, P690, S641, P642, G648, L649, and F647, and they are conserved in all AAVs.

    “The residues forming the nucleotide binding pocket are V418, P419, H639, P690, S641, P642, G648, L649, and F647, and they are conserved in all AAVs.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationreasoning

    Despite conservation of the nucleotide binding pocket, the affinity for the packaged genome is likely reduced in AAV2, resulting in disorder as resolution increases.

    “These observations suggest that despite the conservation of the binding pocket, the affinity for the packaged genome is likely reduced in AAV2 resulting in disorder as resolution increases.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationreasoning

    This implies that rAAV2.7m8 is already primed for endosomal escape or that this capsid region samples at least two conformations.

    “Two of the residues in the pH quartet, Y714 (Y704 in AAV2) and E563, are in the reported AAV8 pH4 conformation (Nam et al., 2011) in rAAV2.7m8, implying that rAAV2.7m8 is already primed for endosomal escape or that this capsid region samples at least two conformations.”
    Quote in the paperSubstantial claim
38 of 38

  • The paper relies on this without support.

    AAV2.7m8 can reach the retina after intravitreal administration, whereas parental AAV2 cannot.

    “For AAV2.7m8, this region is also important for cellular trafficking, given its ability to reach the retina following intravitreal administration, a capability not shared by parental AAV2.”

  • The paper relies on this without support.

    AAV2's 3-fold protrusion matters for receptor binding and antigenicity.

    “In this case, the new vector’s structure retained the overall VP topology and capsid morphology of the parental AAV2 except at the 3-fold protrusion important for receptor binding and antigenicity.”

  • The paper relies on this without support.

    AAV2.7m8 came from a directed evolution screen for viruses with changed tropism compared with the parent.

    “This study utilized cryo-EM to determine the atomic structure of AAV2.7m8, a vector identified from a directed evolution approach to isolate viruses with altered tropism relative to the parental virus.”
3 of 3

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.

  • Conflicting C-terminal residue for rAAV2.7m8

    Minor

    The methods place the last resolved rAAV2.7m8 residue at 736, while the results place it at 745 and identify 745 as the last C-terminal residue.

1 of 1

Cites

Flagged first, then by citations

2 of the 104 references couldn’t be matched to a known paper.

Research works printed but not linked: 1

  1. Afonine, 2013, New tool: phenix.real_space_refine, Comput. Crystallogr. Newslett., 4, 43

Cited by

Sorted by citations

Related

Authors
Bennett, A., Keravala, A., Makal, V., Kurian, J., Belbellaa, B., Aeran, R., Tseng, Y.-S., Sousa, D., Spear, J., Gasmi, M., & Agbandje-McKenna, M.
Affiliations
  • University of Florida (US)
  • Florida State University (US)
Corresponding author
Gasmi, Mehdi
ORCID
Venue
Journal of Structural Biology
Publisher
Elsevier BV
Published
2020-02
Volume
209
Issue
2
Pages
107433
ISSN
1047-8477 (ISSN-L)
DOI
10.1016/j.jsb.2019.107433
PMC
PMC7058502
OpenAlex
W2996414279
Semantic Scholar
ca66ffb43433720d77810b9aeeb88c04a7abdf66
Cited by
33
References
104
Citations by year
2026: 2 · 2025: 8 · 2024: 2 · 2023: 3 · 2022: 5 · 2021: 8 · 2020: 4 · 2015: 1
FWCI
2.56
Type
journal article
Language
en
Open access
Licence
Locations
  • Journal of Structural Biology (publishedVersion)
  • PubMed (publishedVersion)
  • Europe PMC (PubMed Central) (acceptedVersion)
  • PubMed Central (acceptedVersion)
Funders
  • Fayetteville State University (U24 GM116788)
  • National Institutes of Health (R01 GM109524)
  • Florida State University (S10 OD018142-01, U24GM116792)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Virus-based gene therapy research
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Retinal Development and Disorders
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Interference and Gene Delivery
Keywords
capsid structure, amino acid insertion, antigenic determinants, antibody escape, retinal transduction, intravitreal administration, capsid engineering, cryo-electron microscopy
MeSH terms
Dependovirus, Animals, Capsid, Genetic Vectors, Humans, Parvovirinae, Cryoelectron Microscopy, Mice

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 2026-10-01.

Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-01.

Cited sources are checked by gpt-6-sol.

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