Studies on the Nitrate Reductase of Escherichia Coli in the Cell-Free State

Abstract

A method of purifying the nitrate reductase obtained by crushingEscherichia coli cells in a ground glass mill is described, involving elution atpH 9.5 and precipitation into slightly acidified acetone, followed by treatmentwith phosphate buffer ( pH 8.0) and dialysis. Methods of estimating theactivity of the enzyme without recourse to coupling with other enzymes areoutlined, viz.: (a) the oxidation time method; (b) the use of photochemicallyreduced methylene blue as hydrogen donor.