Glyceraldehyde-3-phosphate Dehydrogenase from Human Erythrocytes

Abstract

The physical and catalytic properties of crystalline glyceraldehyde-3-phosphate dehy-drogenase [EC 1. 2.1.12] from human erythrocytes were compared with those of the enzyme from rabbit muscle. The molecular weight of the erythrocyte enzyme was found to be 137, 000 which was close to that of the rabbit muscle enzyme. The two enzymes behaved differently on electrophoresis. The amino acid compositions of the two enzymes were very similar though there were eight half cystines per mole of the erythrocyte enzyme: half as many as in the rabbit enzyme. Four of the sulf-hydryl groups in the enzyme were apparently free and were essential for enzyme activity. The other four SH-groups were hidden but reacted with p-chloromercuri-benzoate when the enzyme was denatured with urea. The erythrocyte enzyme contained less than one mole of NAD, which was bound firmly to the crystallized preparation. However, it was found that the enzyme could bind approximately 3.5 moles of the coenzyme. The enzyme from erythrocytes had essentially the same molecular activity, pH-activity curve and substrate specificity as the rabbit enzyme. Kinetic studies indicated that the reaction of the erythrocyte enzyme was an ordered mechanism of substrate addition. Various kinetic parameters of the reaction are reported.