The purification and properties of histidinol dehydrogenase from Neurospora crassa

Abstract

A procedure is described for the purification of L-histidinol dehydrogenase (L-histidinol-NAD oxidoreductase, EC 1.1.1.23) from Neurospora crassa. The enzyme, as purified, has a sedimentation coefficient, S20, of 7.1 s and a molecular weight of 81000. Considerable variation is possible in the state of polymerization of the enzyme, giving rise to observed molecular weights from 40,000 to 240,000. Several kinetic parameters of the enzyme have been determined. The enzyme is maximally active at pH 9.8; the Km (NAD) is 13.0 x 10-5 M and Km (histidinol) is 8.2 x 10-6 M. The enzyme is highly specific, does not oxidize a range of amino alcohols and other aliphatic alcohols nor reduce NADP and has no demonstrable affinity for histidine. The turnover number is 49 moles of NAD re-duced/min./mole of enzyme (mol. wt. 40,000).