Analysis of a 17-amino acid residue, virus-neutralizing microantibody.
The antibody-binding site, through which an antibody binds to its epitope, is a complex structure formed by the folding together of six complementarity-determining regions (CDRs). However, certain peptides derived from CDR sequences retain antibody specificity and function; these are know as microantibodies (MicroAbs). For example, the F58 MicroAb is a 17 residue, cyclized peptide (CDLIYYDYEEDYYFDYC) derived from CDR-H3 of F58, an IgG1 specific for the gp120 envelope glycoprotein of human immunodeficiency virus type 1 (HIV-1). Both MicroAb and IgG recognize the same epitope in the V3 loop and, despite its small size, the MicroAb neutralizes the infectivity of HIV-1 IIIB only 32-fold less efficiently on a molar basis. The advantage of MicroAbs is that their small size facilitates structure-function analysis. Here, the F58 MicroAb was investigated using alanine scanning, mass spectroscopy and surface plasmon resonance. Neutralization of infectious IIIB was generally more sensitive to alanine substitution than binding to soluble gp120. There appeared to be a division of function within the MicroAb, with some residues involved in antigen binding (alanine substitution of 11D, 12Y or 13Y abrogated both binding and neutralization), whereas others were concerned solely with neutralization (substitution of 3L, 8Y or 14F abrogated neutralization, but not binding). The MicroAb is predominantly beta-sheet and has strong conformational constraints that are probably essential for activity. The MicroAb and soluble gp120 formed a 1 : 1 complex, with an association rate that was threefold greater than that with IgG and a faster dissociation rate. Its equilibrium dissociation constant is 37.5-fold greater than that of IgG, in line with neutralization data. This study demonstrates how MicroAbs can make a useful contribution to the understanding of antigen-antibody interactions.
Heap
Caroline J
CJ
Department of Biological Sciences, University of Warwick, Coventry CV4 7AL, UK.
Wang
Yuqin
Y
Department of Biological Sciences, University of Warwick, Coventry CV4 7AL, UK.
Pinheiro
Teresa J T
TJT
Department of Biological Sciences, University of Warwick, Coventry CV4 7AL, UK.
Reading
Steven A
SA
Department of Biological Sciences, University of Warwick, Coventry CV4 7AL, UK.
Jennings
Keith R
KR
Department of Biological Sciences, University of Warwick, Coventry CV4 7AL, UK.
Dimmock
Nigel J
NJ
Department of Biological Sciences, University of Warwick, Coventry CV4 7AL, UK.
D000409:Alanine; D019943:Amino Acid Substitution; D000918:Antibody Specificity; D000936:Antigen-Antibody Complex / Q000032:analysis; D022801:Complementarity Determining Regions / Q000235:genetics / Q000276:immunology; D000939:Epitopes / Q000276:immunology; D015483:HIV Antibodies / Q000276:immunology*; D015699:HIV Envelope Protein gp120 / Q000235:genetics / Q000276:immunology; D015497:HIV-1 / Q000276:immunology*; D006801:Humans; D013058:Mass Spectrometry; D009500:Neutralization Tests; D009838:Oligodeoxyribonucleotides; D010446:Peptide Fragments / Q000276:immunology; D020349:Surface Plasmon Resonance
C032558:2',5'-oligoadenylate cordycepin analog; D000936:Antigen-Antibody Complex; D022801:Complementarity Determining Regions; D000939:Epitopes; D015483:HIV Antibodies; D015699:HIV Envelope Protein gp120; C059762:HIV envelope protein gp120 (305-321); D009838:Oligodeoxyribonucleotides; D010446:Peptide Fragments; D000409:Alanine