Subunit interactions change the heme active-site geometry in p-cresol methylhydroxylase.
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 cpc and of oxidized p-cresol methylhydroxylase. Analysis of the two-dimensional nuclear Overhauser enhancement and exchange spectroscopy (NOESY) spectrum of reduced cytochrome cpc suggests that the axial ligand, Met-50, of the stable subunit reorients by a rotation about the C gamma-S delta bond when cytochrome cpc binds to the flavin subunit. This reorientation must result in a change in bonding at the heme, which is reflected both in the para-magnetically 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.
McLendon
G L
GL
Department of Chemistry, University of Rochester, NY 14627.
Bagby
S
S
Charman
J A
JA
Driscoll
P C
PC
McIntire
W S
WS
Mathews
F S
FS
Hill
H A
HA
D000494:Allosteric Regulation; D001665:Binding Sites; D003580:Cytochromes / Q000737:chemistry*; D006418:Heme / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D006899:Mixed Function Oxygenases / Q000737:chemistry*; D011487:Protein Conformation; D016958:Pseudomonas putida / Q000201:enzymology
D003580:Cytochromes; D006418:Heme; D006899:Mixed Function Oxygenases; C049502:4-cresol dehydrogenase (hydroxylating)