Characterization of monoclonal antibodies against human prorenin profragment and identification of active prorenins in plasma.
Monoclonal antibodies were raised against a synthetic peptide (43 amino acid residues) that corresponds to the complete profragment of human prorenin. Seven monoclonal antibodies were chosen for further characterization. Two antibodies, 2-X-C1 and 4-X-E1, reacted with the middle region and C-terminus of the profragment and were isotyped IgG1. The affinity constants of these antibodies against the human profragment were 7.6 x 10(8) and 3.0 x 10(7) M-1, respectively. Immunoaffinity columns containing the antibodies 2-X-C1 and 4-X-E1, respectively, were used for the characterization of active prorenin in human plasma. This active prorenin strongly bound to the 4-X-E1 column and eluted as two separate peaks which corresponded to fully and partially active prorenin, respectively. The partially active prorenin had higher activity with a small substrate, tridecapeptide, than with a large one, angiotensinogen, although the fully active prorenin had the same renin activity irrespective of the size of the substrate. These data suggest that new forms of prorenin, active prorenin, exist in human plasma and that their active sites are completely or partially exposed to the substrates. Moreover, the active prorenin in plasma was found not only in human but also in all tested mammalians. Cross-reactivity among the profragments of mammalian plasma prorenins can be explained by conservation of the amino acid sequence (epitope) of the combining site.
Ishizuka
Y
Y
Institute of Applied Biochemistry, University of Tsukuba, Ibaraki.
Iizumi
S
S
Akiyama
F
F
Hori
H
H
Poorman
R A
RA
Lobl
T J
TJ
Murakami
K
K
D000911:Antibodies, Monoclonal / Q000276:immunology*; D000915:Antibody Affinity; D001666:Binding Sites, Antibody; D003429:Cross Reactions; D004792:Enzyme Precursors / Q000097:blood / Q000276:immunology*; D006801:Humans; D015151:Immunoblotting; D009101:Multiple Myeloma / Q000276:immunology; D010446:Peptide Fragments / Q000276:immunology; D012083:Renin / Q000097:blood / Q000276:immunology*
D000911:Antibodies, Monoclonal; D004792:Enzyme Precursors; D010446:Peptide Fragments; D012083:Renin