Extremely thermostable glutamate dehydrogenase from the hyperthermophilic archaebacterium Pyrococcus furiosus

Abstract

The hyperthermophilic archaebacterium Pyrococcus furiosus contains high levels of NAD(P)-dependent glutamate dehydrogenase activity. The enzyme could be involved in the first step of nitrogen metabolism, catalyzing the conversion of 2-oxoglutarate and ammonia to glutamate. The enzyme, purified to homogeneity, is a hexamer of 290 kDa (subunit mass 48 kDa). Isoelectric-focusing analysis of the purified enzyme showed a pI of 4.5. The enzyme shows strict specificity for 2-oxoglutarate and l-glutamate but utilizes both NADH and NADPH as cofactors. The purified enzyme reveals an outstanding thermal stability (the half-life for thermal inactivation at 100°C was 12 h), totally independent of enzyme concentration. P. furiosus glutamate dehydrogenase represents 20% of the total protein; this elevated concentration raises questions about the roles of this enzyme in the metabolism of P. furiosus.