RESUMO
The controversy about the occurrence of an (ADPribosyl)ating activity in yeast is still standing up. Here we discuss this topic on the basis of results obtained with classic experiments proposed over years as basis to characterize an (ADPribosyl)ation system in any organism. Independent results obtained in two different laboratories were in line with each other and went towards the occurrence of an active (ADPribosyl)ating system in Saccharomyces cerevisiae. In fact data collected from nuclear preparations of cultured cells matched those from baker's yeast and lyophilized yeast cells. Yeast (ADPribosyl)ating enzyme is a protein of 80-90 kDa, as determined by electrophoresis on polyacrylamide gel in sodium dodecyl sulphate, followed by immunoblotting with antibodies against anti-poly(ADPribose) polymerase catalytic site. It synthesizes products, that, after digestion with phosphodiesterase, co-migrates mainly with phosphoribosyl adenosine monophosphate after thin layer chromatography on silica gel plate.
Assuntos
Adenosina Difosfato Ribose/metabolismo , Saccharomyces cerevisiae/metabolismo , Autorradiografia , Cromatografia em Camada Fina , Eletroforese em Gel de Poliacrilamida , Proteínas de Saccharomyces cerevisiae/metabolismoRESUMO
The activities of nuclear enzymes involved in NAD+ metabolism in Saccharomyces cerevisiae strain 913a-1 and its mutant 110 previously selected as an NAD+ producer were investigated. The presence of extracellular nicotinamide increased the total NAD+ pool in the cells and increased [3H]nicotinic acid incorporation; however, NAD+ concentration in isolated nuclei decreased slightly. The stimulating effect of nicotinamide on intracellular synthesis of NAD+ correlated with increases in ADP-ribosyl transferase, NAD+-pyrophosphorylase, and NAD+ase activities.