Studies on the charge isomers of superoxide dismutase.

Bovine erythrocycte superoxide dismutase (BRSOD) is a Cu/Zn enzyme that catalyzes the conversion of the superoxide radical to peroxide plus oxygen, thus serving as a scavenger of this reactive and potentially dangerous molecule. The structure of BESOD and the nature of its catalysis have been extensively studied; however, questions remain regarding the regulation of its production within the cell and control of its enzymatic activity. Speculation also surrounds the possible role of naturally occurring change isomers in these processes. The work in this thesis is an attempt to elucidate structural moities that could account for the charge differences in BESOD isomers in order to deduce a possible regulatory role. The experiments can be divided into two main categories: (1) investigation of phosphorylation as a possible modification leading to charge differences, and (2) the nature of the effect of sulfhydryls upon the observed charge isomer distribution. In the first set of experiments BESOD was exposed to a variety of known phosphoproteins phosphatases. It was reasoned that if phosphorylation was the cause of charge isomer occurrence, than dephosphorylation should change the charge isomer distribution pattern. Under the limits of experimental conditions, there was no evidence to suggest that a phosphorylation mechanism could account for charge isomers. In the second set of experiments, BESOD was incubated with beta-mercaptoethanol (beta-ME) and dithiothrietol (DTT). This led to an alteration of the charge isomer patter on gel electrophoresis, with enhancement of the more negatively charged species. Upon removal of beta-ME and DTT from the media, this effect was reversible. It can be speculated that a sulfhydryl end group of BESOD such as cysteine or methionine could account for charge isomers.