ABSTRACT p -Cresol methylhydroxylases (PCMH) from aerobic and facultatively anaerobic bacteria are soluble, periplasmic flavocytochromes that catalyze the first step in biological p -cresol degradation, the hydroxylation of the substrate with water. Recent results suggested that p -cresol degradation in the strictly anaerobic Geobacter metallireducens involves a tightly membrane-bound PCMH complex. In this work, the soluble components of this complex were purified and characterized. The data obtained suggest a molecular mass of 124 ± 15 kDa and a unique αα′β ₂ subunit composition, with α and α′ representing isoforms of the flavin adenine dinucleotide (FAD)-containing subunit and β representing a c -type cytochrome. Fluorescence and mass spectrometric analysis suggested that one FAD was covalently linked to Tyr ³⁹⁴ of the α subunit. In contrast, the α′ subunit did not contain any FAD cofactor and is therefore considered to be catalytically inactive. The UV/visible spectrum was typical for a flavocytochrome with two heme c cofactors and one FAD cofactor. p -Cresol reduced the FAD but only one of the two heme cofactors. PCMH catalyzed both the hydroxylation of p -cresol to p -hydroxybenzyl alcohol and the subsequent oxidation of the latter to p -hydroxybenzaldehyde in the presence of artificial electron acceptors. The very low K _m values (1.7 and 2.7 μM, respectively) suggest that the in vivo function of PCMH is to oxidize both p -cresol and p -hydroxybenzyl alcohol. The latter was a mixed inhibitor of p -cresol oxidation, with inhibition constants of a K _ic (competitive inhibition) value of 18 ± 9 μM and a K _iu (uncompetitive inhibition) value of 235 ± 20 μM. A putative functional model for an unusual PCMH enzyme is presented.