title: Evidence for a methylammonium-binding site on methylamine dehydrogenase of Thiobacillus versutus.
abstract: The nonconvertible substrate analogues di-, tri-, and tetramethylammonium are bound with fairly high affinity to oxidized methylamine dehydrogenase (MADHox) from Thiobacillus versutus and induce the same red-shift in the optical absorbance spectrum of MADHox as do the monovalent cations Cs+, Rb+, and NH4+. Like the monovalent cations, trimethylamine also competitively inhibits the reduction of MADHox by methylamine. Rapid-scan experiments show that within the first few milliseconds of the reaction between MADHox and methylamine a red-shifted intermediate is formed as well. Taken together these experiments demonstrate the existence of a common binding site on MADHox for the substrate CH3NH3+, the substrate analogues (CH3)2NH2+, (CH3)3NH+, and (CH3)4N+, and the monovalent cations Cs+, Rb+, and NH4+. Therefore we conclude that, prior to conversion, methylamine is noncovalently bound to MADHox as a cation. The resonance Raman spectra of MADHox in the absence and presence of Cs+, NH4+, and (CH3)3NH+ are very similar, except for the C=O stretching frequencies of the o-quinone carbonyls of the tryptophyltryptophanquinone (TTQ) active center, which show 5-30 cm-1 downshifts. From these Raman results and the X-ray crystal structure, we conclude that the CH3NH3+ binding site is in close proximity to the O6 carbonyl oxygen of the TTQ.
authors.affiliation: Department of Microbiology and Enzymology, Delft University of Technology, The Netherlands.
authors.forename: A C
authors.initials: AC
authors.lastname: Gorren
authors.forename: P
authors.initials: P
authors.lastname: Moenne-Loccoz
authors.forename: G
authors.initials: G
authors.lastname: Backes
authors.forename: S
authors.initials: S
authors.lastname: de Vries
authors.forename: J
authors.initials: J
authors.lastname: Sanders-Loehr
authors.forename: J A
authors.initials: JA
authors.lastname: Duine
mesh_terms: D001665:Binding Sites; D007700:Kinetics; D008744:Methylamines / Q000737:chemistry*; D010084:Oxidation-Reduction; D000587:Oxidoreductases Acting on CH-NH Group Donors / Q000737:chemistry* / Q000378:metabolism; D013057:Spectrum Analysis; D013059:Spectrum Analysis, Raman; D013855:Thiobacillus / Q000201:enzymology*
chemical_list: D008744:Methylamines; C027451:methylamine; C021567:methylamine dehydrogenase; D000587:Oxidoreductases Acting on CH-NH Group Donors