Photoregulation process of sorghum leaf phosphoenolpyruvate carboxylase: study with monoclonal antibodies

Monoclonal antibodies were produced against the G isozyme subunit of PEP carboxylase (PEPC) from Sorghum leaves by the hybridoma technique. More than 400 antibodies-producing hybridomas to PEPC were produced from the fusion of spleen cells from immunized mice with NS1 myeloma cells. By using an ELISA, three hybridomas (91-G, 83-G, 49-EG) were selected. Monoclonal antibodies were subsequently characterized in a Western experiment; Mabs 83-G and 91-G were found to be highly specific to the G isozyme whereas Mab 49-EG recognized both forms (E and G isozymes) of the enzyme. Addition of Mabs to the enzyme preparation did not modify its catalytic activity nor its activation by glycine. Use of these probes provided direct and definite evidence of the specific enhancing effect of light on the G form and on its corresponding mRNA.