Modification of DNA topoisomerase I enzymatic activity with phosphotyrosyl protein phosphatase and alkaline phosphatase from the hepatopancreas of the shrimp Penaeus japonicus (Crustacea:Decapoda).
DNA topoisomerase I was partially purified from the hepatopancreas of the shrimp Penaeus japonicus. The specific activity of the final preparation was 7,000,000 units/mg of protein with SV40 viral DNA as substrate. SDD-polyacrylamide gel electrophoresis of the final preparation yielded two major bands of proteins with M(r) 70,000 and M(r) 67,000, as well as less intense bands of proteins with M, 64,000 and M(r) 56,000. Incubation of the partially purified enzyme fraction with rabbit antiserum against human DNA topoisomerase I, allowed all these proteins except that of M(r) 56,000, to be positively reacted. Treatment of the partially purified DNA topoisomerase I with tyrosine kinase p43v-abl resulted in phosphorylation of only the two major subunits. Phosphorylation by tyrosine kinase p43v-abl or dephosphorylation by phosphotyrosyl protein phosphatase resulted in a decrease of the enzymatic activity. The treatment with shrimp alkaline phosphatase abolished the enzymatic activity of the purified DNA topoisomerase I in a dose-dependent manner. Thus, the DNA topoisomerase I was apparently isolated from the hepatopancreas of the shrimp P. japonicus in a phosphorylated form, and this phosphorylation was essential for expression of enzymatic activity in vitro. The activity of DNA topoisomerase I is inhibited by ZnCl2, CuCl2 and Pb(NH3)3 at millimolar concentrations, but less inhibition was observed with CaCl2.
Chuang
N N
NN
Division of Biochemistry and Molecular Science, Institute of Zoology, Academia Sinica, Nankang, Taipei, Taiwan.
Lin
C L
CL
Chen
H K
HK
D000469:Alkaline Phosphatase / Q000378:metabolism*; D000818:Animals; D002412:Cations / Q000378:metabolism; D004264:DNA Topoisomerases, Type I / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D004587:Electrophoresis, Agar Gel; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008099:Liver / Q000737:chemistry / Q000201:enzymology; D008670:Metals / Q000378:metabolism; D010179:Pancreas / Q000737:chemistry / Q000201:enzymology; D033561:Penaeidae / Q000201:enzymology*; D010766:Phosphorylation; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism*; D011817:Rabbits; D014443:Tyrosine / Q000378:metabolism
D002412:Cations; D008670:Metals; D014443:Tyrosine; D000469:Alkaline Phosphatase; D017027:Protein Tyrosine Phosphatases; D004264:DNA Topoisomerases, Type I