The biochemical inhibition mode of bredinin-5'-monophosphate on DNA polymerase beta.
We reported previously [T. Horie, Y. Mizushina, M. Takemura, F. Sugawara, A. Matsukage, S. Yoshida, K. Sakaguchi, Int. J. Mol. Med., 1 (1998) 83-90.] that a 5'-monophosphate form (breMP) of bredinin, which has been used clinically as an immunosuppressive drug, selectively suppressed the activities of mammalian DNA polymerase alpha (pol. alpha) and beta (pol. beta). In a preliminary study of the action mode, for pol. beta, breMP acted by competing with, unexpectedly, not only the substrate but also with the template-primer. The mode might be attributable to the structure and function of pol. beta itself. We therefore investigated the biochemical inhibition mode of pol. beta in more detail by using two pol. beta fragments which were proteolytically separated into the template-primer-binding domain and the catalytic domain. BreMP inhibited only the catalytic activity of the catalytic domain fragment, and could not bind to the template-primer-binding domain fragment, suggesting that it directly competes with the substrate at its binding site of the catalytic domain, and indirectly, but simultaneously and competitively disturbs the template-primer incorporation into the template-primer-binding domain.
Mizushina
Y
Y
Department of Applied Biological Science, Science University of Tokyo, Noda, Chiba 278-8510, Japan.
Matsukage
A
A
Sakaguchi
K
K
D000818:Animals; D000970:Antineoplastic Agents / Q000008:administration & dosage / Q000494:pharmacology*; D017104:Bacteriophage M13; D001665:Binding Sites / Q000187:drug effects; D001667:Binding, Competitive; D002384:Catalysis / Q000187:drug effects; D019951:DNA Polymerase beta / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D017931:DNA Primers / Q000737:chemistry / Q000378:metabolism; D004279:DNA, Viral / Q000737:chemistry / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D003848:Deoxyguanine Nucleotides / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000008:administration & dosage / Q000494:pharmacology*; D007700:Kinetics; D008970:Molecular Weight; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D051381:Rats; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors; D012263:Ribonucleosides / Q000008:administration & dosage / Q000494:pharmacology; D012265:Ribonucleotides / Q000008:administration & dosage / Q000494:pharmacology*; D013379:Substrate Specificity
D000970:Antineoplastic Agents; D017931:DNA Primers; D004279:DNA, Viral; D004268:DNA-Binding Proteins; D003848:Deoxyguanine Nucleotides; D004791:Enzyme Inhibitors; D010455:Peptides; D011994:Recombinant Proteins; D012263:Ribonucleosides; D012265:Ribonucleotides; C010052:mizoribine; C062485:bredinin 5'-monophosphate; C029603:deoxyguanosine triphosphate; D019951:DNA Polymerase beta