Immunocytochemical localization of carbamyl phosphate synthetase in the filamentous heterocystous cyanobacteriumNostoc PCC 73102

Summary Free-living nitrogen-fixingNostoc PCC 73102 cells, a filamentous heterocystous cyanobacterium originally isolated from the cycadMacrozamia, were grown without or with the addition of either citrulline or ornithine and examined for the presence of carbamyl phosphate synthetase (CPS) by SDS-PAGE and Western immunoblots. Transmission electron microscopy and immunocytochemical labelling were used to study the cellular and subcellular distribution of CPS in theNostoc cells.Western immunoblots revealed that a polypeptide with a molecular weight of approximately 130 kDa was immunologically related to CPS purified fromE. coli. Nitrogen-fixingNostoc 73102 cultures grown without or with the addition of either citrulline or ornithine showed no differences in their CPS-polypeptide levels, indicating no regulatory effect on the CPS-protein level by these two amino acids. Immunolocalization demonstrated that the CPS protein was located both in vegetative cells and heterocysts, subcellularly evenly distributed over the two cell-types. Using the particle analysis of an image processor and cells grown both without or with addition of either citrulline or ornithine, about 2.5 times more CPS-gold labelling per cell area were observed in the photosynthetic vegetative cells compared to the nitrogen-fixing heterocysts.Abbreviations CPS carbamyl phosphate synthetase - IgG immunoglobulin G - OCT omithine carbamyl transferase