Isoelectrofocusing of rat muscle adenylosuccinase

Adenylosuccinase catalyses the conversion of adenylosuccinic acid to AMP and fumarate. We have developed a coupled enzyme staining procedure applicable to nitrocellulose blots after agarose gel isoelectrofocusing of rat muscle adenylosuccinase. The coupling enzymes, fumarase (fumarate to L-malate) and malic enzyme (L-malate to pyruvate and NADPH), are adsorbed to nitrocellulose prior to blotting. The NADPH, mediated by phenazine methosulfate, converts a tetrazolium salt to its blue formazan. This procedure demonstrated that rat muscle adenylosuccinase consists of three isomeric forms present in similar amounts.