Isoionic titration and isopycnic density gradient centrifugation studies of magnesium activation and subunit dissociation in yeast enolase.

Isopycnic density gradient centrifugations were performed on yeast enolase A in cesium chloride and sulfate at the isoionic pH (6.0–6.5) and at pH 8.1. The dissociation of the enzyme appears to be greater at the more alkaline pH. No large effect of the cofactor magnesium or pH or dissociation on the isopycnic point was found. Isoionic titrations were carried out in the presence and absence of magnesium using both salts. The metal reduces net anion binding by isoionic enzyme and net cation binding by pH 8.1 enzyme by about 1.6 – 3.4 equivalents at 0.05 ionic strength and somewhat less at the isopycnic ionic strengths. It is concluded that the metal does not significantly affect net hydration of the enzyme and that subunit dissociation is not accompanied by large changes in hydration or salt binding.