PURPOSE: To provide a hydroxylase useful for the industrial production of protocatechuic acid.

CONSTITUTION: A reduced nicotinamide adenine dinucleotide (NADH)-dependent p-hydroxybenzoic hydroxylase having the following properties is provided: optimum pH: 7.5-7.8pH using p-hydroxybenzoic acid as a specific substrate; pH 7.4-7.8 as a stable pH range (25°C for 1hr); 33°C as an optimum temperature; stable up to 25°C when incubated at pH7.6 for 60min and completely inactivated at ≥60°C; about 67,000 molecular weight (measured by the high-performance gel permeation column method); inhibited by halide or thiocyanate ions and at least about 10U/mg specific activity (reducing pyridine nucleotide at 340nm). This hydroxylase is capable of acting on p-hydroxybenzoic acid in the presence of the NADH and producing protocatechuic acid and obtained from a culture of a mutant strain of a microorganism, belonging to the genus Rhodococcus and having the ability to produce the NADH-dependent p-hydroxygenzoic hydroxylase, e.g. Rhodococcus L-IE-108 (FERM P-12931).