Bovine thymus poly(adenosine diphosphate ribose) polymerase. Physical properties and binding to DNA.
Purified bovine thymus poly(adenosine diphosphate ribose) polymerase is a monomeric protein with a single polypeptide chain having a molecular weight of approximately 130,000, determined by sodium dodecyl sulfate-gel electrophoresis, analytical ultracentrifugation, and gel filtration. A high frictional ratio (1.81) indicated that the molecule has an elongated shape, or a high solvation, or both. The enzyme is a basic protein (pI 9.8), and amino acid analysis showed a relatively high lysine content. The enzyme activity is dependent on double-stranded DNA and is solely correlated with single- or double-stranded breaks on the DNA. Filter binding assay technique showed that the enzyme-activating efficiency of DNA correlated sufficiently with its enzyme-binding efficiency. Thus, a very high enzyme-activating efficiency of a DNA fraction (active DNA) which was separated from the crude enzyme fraction is mainly due to its high enzyme-binding efficiency. It was also shown that single-stranded DNA and heparin had a strong inhibitory effect on the binding of the enzyme to double-stranded DNA, whereas competitive inhibitors did not affect the binding, We interpret these results to indicate that the binding of the enzyme to double-stranded DNA is a prerequisite step to its catalytic activity and has a dual function: (a) to position the enzyme on specific binding sites such as single- or double-stranded breaks on the DNA, and (b) to induce an active conformation of the enzyme.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D003086:Bacteriocin Plasmids; D001667:Binding, Competitive; D002417:Cattle; D002474:Cell-Free System; D004247:DNA / Q000378:metabolism; D004269:DNA, Bacterial / Q000378:metabolism; D003851:Deoxyribonucleases / Q000378:metabolism; D004926:Escherichia coli; D006358:Hot Temperature; D008970:Molecular Weight; D009244:NAD+ Nucleosidase / Q000032:analysis*; D009691:Nucleic Acid Denaturation; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D011065:Poly(ADP-ribose) Polymerases / Q000032:analysis* / Q000378:metabolism; D012965:Sodium Chloride / Q000494:pharmacology; D013950:Thymus Gland / Q000201:enzymology*
D000596:Amino Acids; D004269:DNA, Bacterial; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D012965:Sodium Chloride; D004247:DNA; D011065:Poly(ADP-ribose) Polymerases; D003851:Deoxyribonucleases; D009244:NAD+ Nucleosidase