14C-labelling of glutamine synthetase and Fe protein of nitrogenase in toluene-treated cells of Rhodopseudomonas capsulata

Abstract

Glutamine synthetase (EC 6.3.1.2) was purified from Rhodopseudomonas capsulata. The molecular weight of the subunit was found to be 56 000 and the isoelectric point of the oligomer 5.5. The subunit protein was localized on discontinuous and two-dimensional polyacrylamide gel electrophoresis patterns. After an ammonia shock produced by addition of 15 mM NH 4Cl to toluene-treated cells preincubated with [ 14C]ATP labelled on adenine, both glutamine synthetase and Fe protein of nitrogenase became inactivated and radioactively labelled, while with [α- 32P]ATP only glutamine synthetase was labelled. Snake venom phosphodiesterase removed the label from glutamine synthetase but not from the Fe protein. This indicates that the binding of the adenine derivative to the Fe protein is not by a phosphate linkage. A glutamine shock, produced by addition of 30 mM glutamine, caused inhibition of nitrogenase and glutamine synthetase activities and 14C labelling of both proteins in the presence of [ 14C]ATP. It is suggested that glutamine may be the active effector in the physiological regulation of nitrogenase.