Biochem Biophys Res Commun. 2016 Nov 11. pii: S0006-291X(16)31847-2. doi: 10.1016/j.bbrc.2016.10.150. [Epub ahead of print]
Streptococcus pneumoniae is a major infectious agent responsible for pneumonia, otitis media, sepsis and meningitis. Pneumococcal surface protein A (PspA) is a well-characterized virulence factor localized on the surface and a target for vaccine development. In this study, we screened a single-chain antibody variable fragment (scFv) using phage display from a human synthetic library to select a clone 2B11. Affinity (Kd) of 2B11 was measured to be 5 nM using biolayer interferometry. 2B11 exhibited a dose-dependent recognition of recombinant PspA with no cross-reactivity towards pneumococcal antigens. The epitope on PspA was defined to residues 231-242 by mutational analysis. Molecular docking analysis supported the experimentally determined epitope, suggesting that the helix spanning residues 231-242 can bind to 2B11 with residues in the CDR-H3 (complementarity determining region 3 in the heavy chain) actively participating in the molecular contacts. Comparison of 2B11 with a commercial PspA antibody revealed that 2B11 exhibited a better specificity towards recombinant PspA antigen. 2B11 was capable of detecting endogenous PspA from pneumococcal lysates with affinity similar to that of the commercial antibody. Our study provides a molecular tool for biosensors detecting pneumococcal diseases.
Copyright © 2016. Published by Elsevier Inc.
2B11; Epitope; Phage display; PspA; Single-chain antibody variable fragment; Streptococcus pneumoniae
PMID: 27845043 DOI: 10.1016/j.bbrc.2016.10.150
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