Sa--a} O Elsevier/North-Holland Biomedical Press CCA 1698 Multiple molecular forms of human cytoplasmic aspartate aminotransferase Robert Rej Dioision oj Lnboratories and Research, New York State Department of Health, Albatxy, NY 12201 (U.S.A.) (Received October 13th, 1980) Summary Human liver cytoplasmic aspartate aminotransferase was found to exhibit five subforms with isoelectric points of 5.15, 5.30, 5.45, 5.60, and 5.80. Treatment with neuraminidase did not affect their electrophoretic mobility. The immunochemical and steady-state kinetic properties of the subforms were identical. Heat treatment increased the proportion of acidic subforms, but all forms were present in fresh tissue. 2-Mercaptoethanol or inhibitors of proteolysis failed to protect against the formation of the subforms with lower isoelectric points. Multiple molecular forms with similar properties svere found for the enzyme of human erythrocytes. This evidence is consistent with deamidation of asparaginyl or glutaminyl residues as the origin of the multiple forms. Human mitochondrial aspartate aminotransferase presented as a single molecular form with an isoelectric point of 9.7. Introduction Aspartate aminotransferase (AspAT, EC 2.6.l.l; t-aspartate:2-oxoglutarate aminotransferase) is present as two genetically and immunologically distinct isoen- zymes in most species [1-5]. One isoenzyme is of mitochondrial origin (m-AspAT); the other is found in the soluble portion of the cell (s-AspAT). Preparations of each isoenzyme have been reported to contain several subforms, which can be detected by their differing electrophoretic mobilities [2-5]. From three to six subforms of s-AspAT have been obtained in various studies [3,6-8]. Marino et al. [7] and Denisova and Polyanovsky [9,10] have published the isolation of five such subforms. The origin of these multiple molecular forms of s-AspAT has been attributed to a number of causes, including oxidation of sulfhydgl groups [11], dearnidation of glutaminyl and asparaginyl residues [6,12,13], conformational changes of the protein [14], differences in pyridoxal phosphate binding [8], and carbohydrate microhetero- geneity [9,10,15]. In the course of purifying human liver s-AspAT [1,16] we have found broad ion-exchange chromatographic elution profiles, which suggested the presence of multiple forms of human s-AspAT with similar but not identical isoelectric points (pI). Since information on the subforms of human s-AspAT is lacking and the nature