Nucleoside diphosphate kinase activity in soluble transducin preparations

摘要: Known nucleoside diphosphate kinases (NDPKs) are oligomers of 17–23-kDa subunits and catalyze the reaction N1TP + N2DP  N1DP + N2TP via formation a histidine-phosphorylated enzyme intermediate. NDPKs involved in activation heterotrimeric GTP-binding proteins (G-proteins) by catalyzing GTP from GDP, but properties G-protein-associated still incompletely known. The aim our present study was to characterize NDPK soluble preparations retinal G-protein transducin. is operationally referred as transducin-NDPK. Like known NDPKs, transducin-NDPK utilizes NTPs phosphorothioate analogs substrates. GDP more effective phosphoryl group acceptor at than ADP CDP, guanosine 5′-[γ-thio]triphosphate (GTP[S]) thiophosphoryl donor adenosine (ATP[S]). In contrast with their action on mastoparan 7 had no stimulatory effect Guanosine 5′-[β,γ-imido]triphosphate (p[NH]ppG) potentiated [3H]GTP[S] [3H]GDP ATP[S] not GTP[S]. Depending donor, [3H]NTP[S] differentially regulated Mg2+, Mn2+, Co2+, Ca2+ Zn2+. [γ-32P]ATP [γ-32P]GTP [32P]phosphorylated, [35S]ATP[S][35S]thiophosphorylated, 36-kDa protein comigrating transducin-β. p[NH]ppG [35S]thiophosphorylation protein. 32P-labeling showed characteristics histidine phosphorylation. There evidence for (thio)phosphorylation proteins. Our data show following: (a) transducin contain GDP-prefering guanine nucleotide-regulated NDPK; (b) transducin-β may serve (thio)phosphorylated intermediate; (c) distinct consist multiple or single kinase regulatory domains.