An Analysis of the Microheterogeneity of PIG Fumarase

Abstract PAGE patterns of pig heart and liver fumarase have been compared in denaturating and non-denaturating conditions. In the presence of 6 M urea, the enzyme, which is fully dissociated and unfolded, migrates as 1 single band at pH 3.2. At pH 3.3, the enzyme, which is dissociated but not extensively unfolded, again migrates as 1 single band. At pH 7.7-9.3 bands of different mobilities are observed when the electrode buffer contains ions which have a specific affinity for the enzyme such as glycine or citrate. IEF in the presence of 4.2 M acetamide reveals essentially 1 band (pI 7.8), whereas in 5 M urea at least 4 main bands are seen; these must be due to carbamylation. IEF in non-denaturating conditions gives rise to a very complex and smeared-out pattern of bands. The microheterogeneity of pig fumarase detected in certain conditions can be explained by a specific binding of ligands from the buffer solutions onto the enzyme.