The Isolation and Characterization of the Isoenzymes of Rabbit‐Muscle Triosephosphate Isomerase

Crystalline and specific‐activity‐constant rabbit‐muscle triosephosphate isomerase was resolved into three major forms, α, β and γ (97%) and two minor components δ and ɛ (3%) through DEAE‐cellulose chromatography. Hybridization studies have shown the α and γ isoenzymes to be homodimers AA and BB, respectively, and β the heterodimer AB. Reassociation studies from 8 M urea have indicated that α, β and γ are not conformers. The evidence supports earlier data suggesting that the heterogeneity of triosephosphate isomerase is largely due to the random combination of two chemically non‐identical subunits (A and B) to give three native dimeric forms. Following hybridization, the pattern obtained with δ and ɛ is more complex. Reassociation of either form resulted in the formation of all five bands as well as two or four, respectively, additional more anodic bands. Substrate and inhibition characterization have shown the isoenzymes to be “isokinetic”. Titration with 5,5′‐dithiobis(2‐nitrobenzoate) has indicated two accessible sulfhydryl groups in the native molecule, one of which appears to be essential for activity.