The green alga Scenedesmus obliquus contains both diadenosine 5',5'''-P1,P4-Tetraphosphate (asymmetrical) pyrophosphohydrolase and phosphorylase activities

Abstract

Diadenosine 5',5'''-P1,P4-tetraphosphate (Ap4A) phosphorylase and Ap4A pyrophosphohydrolase activities have been purified from extracts of the green alga Scenedesmus obliquus. Both activities were also detected in Scenedesmus basiliensis, Scenedesmus quadricauda and in Chlorella vulgaris. This is the first time that both types of enzyme have been detected in the same species. The Ap4A phosphorylase has a molecular mass of 46-48 kDa, a broad pH optimum between 7.5 and .5, and requires a divalent ion for activity (Mg2+ > Co2+ > Ca2+ = Mn2+ = d2+ > Zn2+). It degrades substrates with at least four phosphate groups and always produces a nucleoside 5'-diphosphate product. The K(m) values for Ap4A and Pi are 5.3μM and 160μM, respectively, and k(cat)= 1.8 s-1. Arsenate, vanadate, molybdate, chromate and tungstate can substitute for phosphate. The enzyme also catalyses Ap4A synthesis (K(eq) = [Ap4A]] [Pi]/[ATP][ADP] = 9 x 10-4) and ADP arsenolysis. The Ap4A hydrolase has a molecular mass of 26-28 kDa, an alkaline pH optimum of 8.8-9.8, and prefers Zn2+ as the stimulatory ion (Zn2+ > Mg2+ > Mn2+ > Co2+ > Cd2+). It degrades substrates with at least four phosphate groups, having a slight preference for Ap5A, and always produces a nucleoside 5'-triphosphate product. The Km value for Ap4A is 6.6 μM and k (cat) = 1.3 s-1, It is inhibited competitively by adenosine 5'-tetraphosphate (K(i)= 0.67μM) and non-competitively by fluoride (K(i) = 150 μM). A 50-54 kDa dinucleoside 5',5'''-P1,P3-triphosphate (Ap3A) pyrophosphohydrolase was also detected in S. obliquus S. quadricauda and C. vulgaris. The corresponding enzyme in S. basiliensis (> 100 kDa) may be a dimer.