Differential changes in ornithine aminotransferase self-affinity produced by exposure to basic amino acids and increases in the intrinsic electronegativity of the enzyme monomer

Abstract In a previous study we showed that ornithine aminotransferase (OAT) exhibits concentration-dependent self-association in two stages (45,000 M r monomers aggregate to form 140,000–150,000 M r trimers; the trimers then aggregate to form higher-molecular-weight complexes). In an attempt to characterize further the molecular mechanisms involved in OAT aggregation, the present study examined the effects of basic amino acids and keto acids on the aggregation process. These experiments showed that basic amino acids (ornithine and lysine) inhibit the association of monomers to form trimers, apparently by interaction with carboxyl groups on the surfaces of the monomers. The aggregation of trimers to form higher-molecular-weight assemblies is not affected by basic amino acids, and neither aggregation stage is affected by the keto acids, α-ketoglutarate, or oxaloacetate. We also found that two different OAT preparations (one fresh, the other 18 months old) differed in aggregation characteristics; the older preparation showed reduced self-affinity at both aggregation stages, but both preparations had similar catalytic efficiencies. Electrophoretic studies indicated that the older preparation contained variants of the enzyme monomer with greater electronegativity than did the fresh preparation. We conclude, therefore, that OAT purification exposes ionically labile but catalytically insignificant domains on the monomer surface, and the loss of positively charged groups from such regions diminishes the OAT aggregation potential.