1. A mutant of Neurospora crassa has been isolated whose pyruvate kinase is twice as active as the wild type enzyme. 2. The purified mutant and the wild type enzymes exhibit similar immunological properties, pI values (6.4) and Arrhenius activation energy (11.2 kcal/mol). 3. Both the enzymes show hyperbolic saturation kinetics with ADP and sigmoidal kinetics with PEP. 4. The mutant enzyme displays a higher affinity for PEP and a greater extent of cooperativity in binding than the wild type. 5. Conformational alterations in the mutant enzyme are inferred on the basis of electrophoretic analyses and denaturation by urea, SDS and heat.