Pumpkin (Cucurbita sp.) seed globulin I. Purification, characterization, and subunit structure

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Abstract
A heat stable globulin present in the cotyledons of pumpkin seeds was prepared as crystals which were soluble in a dilute saline solution below pH 4.5 or in a solution with a high ionic strength at neutral pHs. The protein was nearly homogeneous by ultracentrifuge analysis, and had a molecular weight of about 112,000 daltons. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis separated the globulin into two subunits, α and β, corresponding to molecular weights of about 63,000 and 56,000 daltons, respectively. By reduction of disulfide bonds, the two subunits were each separated into two polypeptide chains with molecular weights of around 36,000 and 22,000 daltons, judged by gel electrophoresis. The amino acid composition of whole globulin indicated high contents of arginine, glutamic acid and aspartic acid. The total number of half-cystine residue was nine and only one residue was shown to be free. The subunit structure of the globulin is discussed. The protein has been shown to have oxaloacetate decarboxylase activity, and this fact was confirmed. However, the activity decreased markedly at pH 4.5 in a fairly short period. It did not require Mn++, and the Km for oxaloacetate was determined to be 4.1 mM.