Specific HIV gp120-cleaving antibodies induced by covalently reactive analog of gp120.
We report the results of efforts to strengthen and direct the natural nucleophilic activity of antibodies (Abs) for the purpose of specific cleavage of the human immunodeficiency virus-1 coat protein gp120. Phosphonate diester groups previously reported to form a covalent bond with the active site nucleophile of serine proteases (Paul, S., Tramontano, A., Gololobov, G., Zhou, Y. X., Taguchi, H., Karle, S., Nishiyama, Y., Planque, S., and George, S. (2001) J. Biol. Chem. 276, 28314-28320) were placed on Lys side chains of gp120. Seven monoclonal Abs raised by immunization with the covalently reactive analog of gp120 displayed irreversible binding to this compound (binding resistant to dissociation with the denaturant SDS). Catalytic cleavage of biotinylated gp120 by three monoclonal antibodies was observed. No cleavage of albumin and the extracellular domain of the epidermal growth factor receptor was detected. Cleavage of model peptide substrates occurred on the C-terminal side of basic amino acids, and Km for this reaction was approximately 200-fold greater than that for gp120 cleavage, indicating Ab specialization for the gp120 substrate. A hapten phosphonate diester devoid of gp120 inhibited the catalytic activity with exceptional potency, confirming that the reaction proceeds via a serine protease mechanism. Irreversible binding of the hapten phosphonate diester by polyclonal IgG from mice immunized with gp120 covalently reactive analog was increased compared with similar preparations from animals immunized with control gp120, indicating induction of Ab nucleophilicity. These findings suggest the feasibility of raising antigen-specific proteolytic antibodies on demand by covalent immunization.
Paul
Sudhir
S
Chemical Immunology Research Center, Department of Pathology, University of Texas, Houston Medical School, 77030, USA. Sudhir.Paul@uth.tmc.edu
Planque
Stephanie
S
Zhou
Yong-Xin
YX
Taguchi
Hiroaki
H
Bhatia
Gita
G
Karle
Sangeeta
S
Hanson
Carl
C
Nishiyama
Yasuhiro
Y
D000595:Amino Acid Sequence; D000818:Animals; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D015483:HIV Antibodies / Q000096:biosynthesis* / Q000276:immunology; D015699:HIV Envelope Protein gp120 / Q000737:chemistry / Q000276:immunology*; D006868:Hydrolysis; D051379:Mice; D021241:Spectrometry, Mass, Electrospray Ionization
D015483:HIV Antibodies; D015699:HIV Envelope Protein gp120