Crossed affinity immunoelectrophoresis of the Escherichia coli pyruvate dehydrogenase complex.

The crossed immunoelectrophoretic pattern obtained with the intact pyruvate dehydrogenase complex of Escherichia coli can be modified when this technique is combined with affinity gel electrophoresis using reactive dyes coupled to agarose as ligands. The patterns that arise have been interpreted with respect to localization of the three component enzymes. This was realized by using antibodies with different specificity, active enzyme staining and E2-E3 subcomplex behaviour. Dissociation of E1 subunits occurs more easily than that of E3 but remains incomplete in this system. The free reactive Procion Blue-MX dyes tested inactivate the complex even at neutral pH. The dyes react with all three components but E3 (80%) and E2 (15-20%) retain part of their catalytic activity. Modification leads to an enhanced dissociation of E1.