Conformational analysis of human serum albumin and its non-enzymatic glycation products using monoclonal antibodies.
Monoclonal antibodies (mAbs) were prepared to analyse the conformation of human serum albumin (HSA) and its non-enzymatic glycation (NEG) products. We first determined the epitopes of the mAbs using HSA subdomains expressed on the surface of yeast. Each mAb was classified as belonging to one of two groups; Type I mAbs which recognized a single subdomain structure and Type II mAbs which bound to plural subdomains. We analysed the pH-dependent conformational change in HSA. We found that one Type II mAb, HAy2, detected the normal to base form (N-B) transition while the other did not, suggesting that N-B transition occurred around Domain I accompanied by topological isomerization of subdomains without changing the subdomain structure itself. Next, we analysed the conformations of the NEG products. Since all mAbs reacted with the early NEG products, no structural change was thought to have occurred in the early NEG products. On the other hand, only a Type I mAb, HAy1, had full binding activity with the advanced glycation end products (AGE) while the other mAbs had lost or had diminished activity, suggesting that the over-all tertiary structure of HSA was altered except for a subdomain, sDOM Ia, in AGE.
Research Institute for Biological Sciences, Tokyo University of Science, 2669, Yamazaki, Noda, Chiba 278-0022, Japan.
Keigo
K
Saito
Kuniko
K
Hamano
Masatoshi
M
Nakagawa
Keiko
K
Yugawa
Jin
J
Muraoka
Hiroyoshi
H
Kuba
Koji
K
Furukawa
Takachika
T
Azuma
D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism*; D000939:Epitopes / Q000737:chemistry / Q000378:metabolism; D017127:Glycation End Products, Advanced / Q000737:chemistry / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D006863:Hydrogen-Ion Concentration; D051379:Mice; D008807:Mice, Inbred BALB C; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D020033:Protein Isoforms / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D012709:Serum Albumin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D013816:Thermodynamics
D000911:Antibodies, Monoclonal; D000939:Epitopes; D017127:Glycation End Products, Advanced; D020033:Protein Isoforms; D012709:Serum Albumin