Purification of three distinct enolase isoenzymes from yeast

A method for the rapid isolation of yeast enolases, yielding three distinct isoenzymes, has been devised. In the first step anionic proteins were precipitated with polyethyleneimine, whereas hydrophobic enolase isoenzymes remained in the supernatant. Secondly, the supernatant was 45% saturated with ammonium sulfate and bound to phenyl-Sepharose CL-4B. Decreasing ammonium sulfate and simultaneously increasing ethylene glycol concentrations were used for elution. Finally, enolase isoenzymes were separated by chromatofocusing. The purified isoenzymes gave single bands after isoelectric focusing.