Subunit Interactions Change the Heme Active-Site Geometry in p-Cresol Methylhydroxylase

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 88; no. 21; pp. 9463 - 9467
• Main Authors: McLendon, George L., Bagby, Stefan, Charman, Judith A., Driscoll, Paul C., McIntire, W. S., Mathews, F. S., H. Allen O. Hill
• Format: Journal Article
• Language: English
• Published: Washington, DC National Academy of Sciences of the United States of America 01.11.1991; National Acad Sciences
• Subjects: Allosteric Regulation; Analytical, structural and metabolic biochemistry; Binding Sites; Biochemistry; Biological and medical sciences; Crystal structure; Cytochromes; Cytochromes - chemistry; Electron density; Enzymes; Enzymes and enzyme inhibitors; Fundamental and applied biological sciences. Psychology; heme; Heme - chemistry; Ligands; Magnetic Resonance Spectroscopy; Mixed Function Oxygenases - chemistry; Oxidoreductases; Protein Conformation; Protons; Pseudomonas putida; Pseudomonas putida - enzymology; Reactivity; Retraining; Spectrometers; Cytochrome; Isoelectrical focusing; Enzyme; Active site; 4-Cresol dehydrogenase(hydroxylating); Molecular reorientation; NMR spectrometry; Subunit
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.88.21.9463

The enzyme p-cresol methylhydroxylase [4-cresol:(acceptor) oxidoreductase (methyl-hydroxylating), EC 1.17.99.1] contains two subunits: a cytochrome c (electron transfer) subunit (cytochrome cpc) and a flavin (catalytic) subunit. When these subunits are separated by isoelectric focusing, a stable cytochrome subunit is obtained. Significant differences are observed between the one-dimensional NMR spectra of oxidized cytochrome cpcand of oxidized p-cresol methylhydroxylase. Analysis of the two-dimensional nuclear Overhauser enhancement and exchange spectroscopy (NOESY) spectrum of reduced cytochrome cpcsuggests that the axial ligand, Met-50, of the stable subunit reorients by a rotation about the Cγ-Sδ bond when cytochrome cpcbinds to the flavin subunit. This reorientation must result in a change in bonding at the heme, which is reflected both in the paramagnetically shifted resonances and in the redox potential. p-Cresol methylhydroxylase thereby provides an interesting example of the coupling of subunit interactions to active-site structure and reactivity.