Intramolecular H-Bonds Govern the Recognition of a Flexible Peptide by an Antibody

Miyanabe, Kazuhiro; Akiba, Hiroki; Kuroda, Daisuke; Nakakido, Makoto; Kusano-Arai, Osamu; Iwanari, Hiroko; Hamakubo, Takao; Caaveiro, J.M.M.; Tsumoto, Kouhei

doi:10.2139/ssrn.3155851

Cited by 6 publications

(10 citation statements)

References 51 publications

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“…We observed the same when evaluating the S2H97, EY6A and S304 antibodies. H-bonds are important non-covalent interaction forces which can assist in protein residue bindings, especially in stabilizing antibody-antigen interactions [26]. Mutated Q380 residue leads to an H-bond disruption observed previously between the wild Y380 with the S99 residue of the CR3022 neutralizing antibody.…”

Section: Resultsmentioning

confidence: 87%

Y380Q novel mutation in receptor-binding domain of SARS-CoV-2 spike protein together with C379W interfere in the neutralizing antibodies interaction

Sartor

Varela

Meireles

et al. 2021

Preprint

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Background: The emergence of SARS-CoV-2 variants is a current public health concern possibly impacting COVID-19 disease diagnosis, transmission patterns and vaccine effectiveness. Objectives: To describe the SARS-CoV-2 lineages circulating early pandemic among samples with S gene dropout and characterize a novel mutation in receptor-binding domain (RBD) of viral spike protein. Study design: Adults and children older than 2 months with signs and symptoms of COVID-19 were prospectively enrolled from May to October 2020 in Porto Alegre, Brazil. All participants performed RT-PCR assays for diagnosing SARS-CoV-2, samples with S gene dropout and Ct < 30 (cycle threshold) were submitted to whole genome sequencing (WGS), and homology modeling and physicochemical properties analysis were performed. Results: 484/1,557 participants tested positive for SARS-CoV-2. The S gene dropout was detected in 7.4% (36/484) as early as May, and a peak was observed in early August. WGS was performed in 8 samples. The B.1.1.28, B.1.91 and B.1.1.33 lineages were circulating in early pandemic. The RBD novel mutation (Y380Q) was found in one sample occurring simultaneously with C379W and V395A, and the B.1.91 lineage in the spike protein. Conclusion: Mutations in the SARS-CoV-2 spike region were detected early in the COVID-19 pandemic in Southern Brazil, regarding the B.1.1.28, B.1.91 and B.1.1.33 lineages identified. The novel mutation (Y380Q) with C379W, modifies important RBD properties, which may interfere with the binding of neutralizing antibodies (CR3022, EY6A, H014, S304).

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Section: Resultsmentioning

confidence: 87%

Y380Q novel mutation in receptor-binding domain of SARS-CoV-2 spike protein together with C379W interfere in the neutralizing antibodies interaction

Sartor

Varela

Meireles

et al. 2021

Preprint

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“…(B) Schematic of NP-SpyCatcher-scFv Ab conjugates. Core/shell nanocrystals (red and dark gray), encapsulated in amphiphilic polymers (light gray), conjugated to a single-Cys SpyCatcher (PDB file 4MLI) 23 and SpyTag-scFv (PDB file 5YD5) 24 chimera. The SpyTag peptide is added at the scFv C terminus, marked (*) in the structure.…”

Section: Resultsmentioning

confidence: 99%

“…Ab engineering efforts over the last 2 decades have demonstrated that selective and high-affinity binding sequences against a variety of antigens can be selected and then subcloned into Ab scaffolds to create stable Ab with non-natural properties and that this requires only basic recombinant DNA technology. 3 Unlike naturally occurring IgG Ab, which are >15 nm along their longest axes, 6 scFv Ab are 2−3 nm in diameter, 24 can readily be expressed in E. coli, and do not have Fc domains that bind to macrophages, lymphocytes, and other cells expressing Fc receptors. 59 A key advantage of recombinant Ab technology is the ease of introducing C-terminal modifications for various applications, ranging from purification (e.g., hexahistidine, myc tags), to valency engineering, to biotechnology applications such as drug conjugation and enzyme fusion.…”

Section: Resultsmentioning

confidence: 99%

Immunotargeting of Nanocrystals by SpyCatcher Conjugation of Engineered Antibodies

et al. 2021

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Inorganic nanocrystals such as quantum dots (QDs) and upconverting nanoparticles (UCNPs) are uniquely suited for quantitative live-cell imaging and are typically functionalized with ligands to study specific receptors or cellular targets. Antibodies (Ab) are among the most useful targeting reagents owing to their high affinities and specificities, but common nanocrystal labeling methods may orient Ab incorrectly, be reversible or denaturing, or lead to Ab-NP complexes too large for some applications. Here, we show that SpyCatcher proteins, which bind and spontaneously form covalent isopeptide bonds with cognate SpyTag peptides, can conjugate engineered Ab to nanoparticle surfaces with control over stability, orientation, and stoichiometry. Compact SpyCatcher-functionalized QDs and UCNPs may be labeled with short-chain variable fragment Ab (scFv) engineered to bind urokinase-type plasminogen activator receptors (uPAR) that are overexpressed in many human cancers. Confocal imaging of anti-uPAR scFv-QD conjugates shows the Ab mediates specific binding and internalization by breast cancer cells expressing uPAR. Time-lapse imaging of photostable scFv-UCNP conjugates show that Ab binding causes uPAR internalization with a ∼20-minute half-life on the cell surface, and uPAR is internalized to endolysosomal compartments distinct from general membrane stains and without significant recycling to the cell surface. The controlled and stable conjugation of engineered Ab to NPs enables targeting of diverse receptors for live-cell study of their distribution, trafficking, and physiology.

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“…Evidence has accumulated that peptides bind to their target proteins by "conformational selection" [51][52][53][54][55], motivating an approach to peptide-protein docking based on pregenerating peptide conformations that are metastable in solution [13]. In order for this pregeneration to yield a complete ensemble and yet occur in a reasonable time, the peptide may need to be represented by a low-resolution model.…”

Section: Discussionmentioning

confidence: 99%

Receptor-free Discrimination of Peptide Native Bound Conformations Suggests Limited Transferability of the “cen_std+score4l” Rosetta Energy Function From Proteins to Peptides

Bazzoli¹,

Contini²

2020

Preprint

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One of the strategies of peptide–protein docking is to pregenerate an ensemble of peptide conformations in the absence of the receptor, and then dock them as rigid bodies onto its surface. Success of this strategy requires that the scoring function that drives the pregeneration step be able to discriminate in favor of conformations that resemble the native bound conformation. Here we present a study on the discrimination of peptide native bound conformations as achieved without receptor by the “cen_std+score4L” Rosetta energy function, a low-resolution scoring function equivalent to one chosen for other tasks where the modeling of solvent effects is of special importance. The cen_std+score4L function was able to assign, on average, lower energies to native-like than to non-native decoy conformations for only 3 of our 18 test peptides; it also ranked one or more native-like decoys in the top 1% for only 2 peptides. However, by optimizing the weights of the energy terms that define the cen_std+score4L function, native discrimination improved substantially: Native-like decoys were assigned lower energies than non-native decoys for 16 peptides, with a discrimination signal larger than noise for 9 peptides, that is, 50% of the test set. And for 9 peptides, too, native-like decoys ranked in the top 1%. An ensuing energetic analysis of native-like versus non-native decoys suggests that native peptide conformations have solvation and non-local electrostatics that poorly recapitulate those of native protein conformations. Native peptide conformations are also characterized by few backbone–backbone H-bonds and by lack of compactness, presumably to optimize interaction with the receptor. Overall, this study lays groundwork for pregenerating dockable peptide conformations with Rosetta, whether the subsequent docking will be performed by Rosetta or some other software.

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Intramolecular H-Bonds Govern the Recognition of a Flexible Peptide by an Antibody

Cited by 6 publications

References 51 publications

Y380Q novel mutation in receptor-binding domain of SARS-CoV-2 spike protein together with C379W interfere in the neutralizing antibodies interaction

Y380Q novel mutation in receptor-binding domain of SARS-CoV-2 spike protein together with C379W interfere in the neutralizing antibodies interaction

Immunotargeting of Nanocrystals by SpyCatcher Conjugation of Engineered Antibodies

Receptor-free Discrimination of Peptide Native Bound Conformations Suggests Limited Transferability of the “cen_std+score4l” Rosetta Energy Function From Proteins to Peptides

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