title: A pentacyclic reaction intermediate of riboflavin synthase.
abstract: The S41A mutant of riboflavin synthase from Escherichia coli catalyzes the formation of riboflavin from 6,7-dimethyl-8-ribityllumazine at a very low rate. Quenching of presteady-state reaction mixtures with trifluoroacetic acid afforded a compound with an absorption maximum at 412 nm (pH 1.0) that can be converted to a mixture of riboflavin and 6,7-dimethyl-8-ribityllumazine by treatment with wild-type riboflavin synthase. The compound was shown to qualify as a kinetically competent intermediate of the riboflavin synthase-catalyzed reaction. Multinuclear NMR spectroscopy, using various 13C- and 15N-labeled samples, revealed a pentacyclic structure arising by dimerization of 6,7-dimethyl-8-ribityllumazine. Enzyme-catalyzed fragmentation of this compound under formation of riboflavin can occur easily by a sequence of two elimination reactions.
authors.affiliation: Lehrstuhl für Organische Chemie und Biochemie, Technische Universität München, Lichtenbergstrasse 4, D-85747 Garching, Germany.
authors.forename: B
authors.initials: B
authors.lastname: Illarionov
authors.forename: W
authors.initials: W
authors.lastname: Eisenreich
authors.forename: A
authors.initials: A
authors.lastname: Bacher
mesh_terms: D019943:Amino Acid Substitution; D002247:Carbon Isotopes; D004926:Escherichia coli / Q000201:enzymology; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D015394:Molecular Structure; D016297:Mutagenesis, Site-Directed; D009587:Nitrogen Isotopes; D011621:Pteridines / Q000737:chemistry* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012256:Riboflavin / Q000096:biosynthesis / Q000737:chemistry; D012258:Riboflavin Synthase / Q000737:chemistry* / Q000378:metabolism*; D013053:Spectrophotometry
chemical_list: D002247:Carbon Isotopes; D009587:Nitrogen Isotopes; D011621:Pteridines; D011994:Recombinant Proteins; C001061:6,7-dimethyl-8-ribityllumazine; D012258:Riboflavin Synthase; D012256:Riboflavin