title: Purification and properties of 5,10-methylenetetrahydromethanopterin dehydrogenase and 5,10-methylenetetrahydromethanopterin reductase, two coenzyme F420-dependent enzymes, from Methanosarcina barkeri.
abstract: 5,10-Methylenetetrahydromethanopterin dehydrogenase and 5,10-methylenetetrahydromethanopterin reductase have been purified to homogeneity by a factor of 86 and 68, respectively, from methanol-grown Methanosarcina barkeri cells. The dehydrogenase was isolated as a hexamer of a single 35 kDa subunit, whereas the reductase was composed of four identical 38 kDa subunits. The purified oxygen-stable enzymes catalyzed the oxidation of 5,10-methylenetetrahydromethanopterin and methyltetrahydromethanopterin with Vmax values of 3000 and 200 mumol min-1 mg-1, respectively. The methanogenic electron carrier coenzyme F420 was a specific electron acceptor for both enzymes. Steady state kinetics for the two enzymes were in agreement with ternary complex (sequential) mechanisms. Methylene reductase and methylene dehydrogenase are proposed to function in the methanol oxidation step to CO2.
authors.affiliation: Department of Microbiology, Faculty of Science, University of Nijmegen, The Netherlands.
authors.forename: B W
authors.initials: BW
authors.lastname: te Brömmelstroet
authors.forename: W J
authors.initials: WJ
authors.lastname: Geerts
authors.forename: J T
authors.initials: JT
authors.lastname: Keltjens
authors.forename: C
authors.initials: C
authors.lastname: van der Drift
authors.forename: G D
authors.initials: GD
authors.lastname: Vogels
mesh_terms: D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D007700:Kinetics; D046911:Macromolecular Substances; D017021:Methanosarcina barkeri / Q000201:enzymology*; D008970:Molecular Weight; D000587:Oxidoreductases Acting on CH-NH Group Donors / Q000302:isolation & purification* / Q000378:metabolism; D012256:Riboflavin / Q000031:analogs & derivatives* / Q000378:metabolism; D013045:Species Specificity; D013056:Spectrophotometry, Ultraviolet; D013816:Thermodynamics
chemical_list: D046911:Macromolecular Substances; C015084:coenzyme F420; D000587:Oxidoreductases Acting on CH-NH Group Donors; C047126:methylenetetrahydromethanopterin dehydrogenase; D012256:Riboflavin