Immunochemical and biological characterization of monoclonal antibodies against BaP1, a metalloproteinase from Bothrops asper snake venom.
BaP1 is a P-I class of Snake Venom Metalloproteinase (SVMP) relevant in the local tissue damage associated with envenomations by Bothrops asper, a medically-important species in Central America and parts of South America. Six monoclonal antibodies (MoAb) against BaP1 (MABaP1) were produced and characterized regarding their isotype, dissociation constant (K(d)), specificity and ability to neutralize BaP1-induced hemorrhagic and proteolytic activity. Two MABaP1 are IgM, three are IgG1 and one is IgG2b. The K(d)s of IgG MoAbs were in the nM range. All IgG MoAbs recognized conformational epitopes of BaP1 and B. asper venom components but failed to recognize venoms from 27 species of Viperidae, Colubridae and Elapidae families. Clone 7 cross-reacted with three P-I SVMPs tested (moojeni protease, insularinase and neuwiedase). BaP1-induced hemorrhage was totally neutralized by clones 3, 6 and 8 but not by clone 7. Inhibition of BaP1 enzymatic activity on a synthetic substrate by MABaP1 was totally achieved by clones 3 and 6, and partially by clone 8, but not by clone 7. In conclusion, these neutralizing MoAbs against BaP1 may become important tools to understand structure-function relationships of BaP1 and the role of P-I class SVMP in snakebite envenomation.
Fernandes
I
I
Laboratório de Imunopatologia, Instituto Butantan, Av. Vital Brazil, 1500, Butantã, CEP 05503-900, São Paulo, SP, Brazil.
Assumpção
G G
GG
Silveira
C R F
CR
Faquim-Mauro
E L
EL
Tanjoni
I
I
Carmona
A K
AK
Alves
M F M
MF
Takehara
H A
HA
Rucavado
A
A
Ramos
O H P
OH
Moura-da-Silva
A M
AM
Gutiérrez
J M
JM
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000276:immunology* / Q000302:isolation & purification; D017837:Bothrops / Q000502:physiology*; D003429:Cross Reactions; D003435:Crotalid Venoms / Q000037:antagonists & inhibitors / Q000201:enzymology* / Q000633:toxicity; D004487:Edema / Q000139:chemically induced / Q000517:prevention & control; D004797:Enzyme-Linked Immunosorbent Assay; D018604:Epitope Mapping; D006470:Hemorrhage / Q000139:chemically induced / Q000517:prevention & control; D015151:Immunoblotting; D007136:Immunoglobulins; D007274:Injections, Intraperitoneal; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors / Q000276:immunology* / Q000633:toxicity; D051379:Mice; D008807:Mice, Inbred BALB C; D009500:Neutralization Tests
D000911:Antibodies, Monoclonal; D003435:Crotalid Venoms; D007136:Immunoglobulins; C104933:BaP1 metalloproteinase; D008666:Metalloendopeptidases