KLEIN, AR and KOCH, J and STETTER, KO and THAUER, RK (1993) 2 N5,N10-METHYLENETETRAHYDROMETHANOPTERIN DEHYDROGENASES IN THE EXTREME THERMOPHILE METHANOPYRUS-KANDLERI - CHARACTERIZATION OF THE COENZYME-F(420)-DEPENDENT ENZYME. ARCHIVES OF MICROBIOLOGY, 160 (3). pp. 186-192. ISSN 0302-8933,
Full text not available from this repository.Abstract
It was recently reported that the extreme thermophile Methanopyrus kandleri contains only a H-2-forming N5,N-10-methylenetetrahydromethanopterin dehydrogenase which uses protons as electron acceptor. We describe here the presence in this Archaeon of a second N5,N-10-methylenetetrahydromethanopterin dehydrogenase which is coenzyme F420-dependent. This enzyme was purified and characterized. The enzyme was colourless, had an apparent molecular mass of 300 kDa, an isoelectric point of 3.7 +/- 0.2 and was composed of only one type of subunit of apparent molecular mass of 36 kDa. The enzyme activity increased to an optimum with increasing salt concentrations. Optimal salt concentrations were e.g. 2 M (NH4)2SO4, 2 M Na2HPO4, 1.5 M K2HPO4, and 2 M NaCl. In the absence of salts the enzyme exhibited almost no activity. The salts affected mainly the V(max) rather than the K(m) of the enzyme. The catalytic mechanism of the dehydrogenase was determined to be of the ternary complex type, in agreement with the finding that the enzyme lacked a chromophoric prosthetic group. In the presence of 1 M (NH4)2SO4 the V(max) was 4000 U/mg (k(cat) = 2400 s-1) and the K(m) for N5,N-10-methylenetetrahydromethanopterin and for coenzyme F420 were 80 muM and 20 muM, respectively. The enzyme was relatively heat-stable and lost no activity when incubated anaerobically in 50 MM K2HPO4 at 90-degrees-C for one hour. The N-terminal amino acid sequence was found to be similar to that of the F420-dependent N5,N-10-methylenetetrahydromethanopterin dehydrogenase from Methanobacterium thermoautotrophicum, Methanosarcina barkeri, and Archaeoglobus fulgidus.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | METHANOBACTERIUM-THERMOAUTOTROPHICUM; N-10-METHYLENETETRAHYDROMETHANOPTERIN DEHYDROGENASE; METHYLENETETRAHYDROMETHANOPTERIN DEHYDROGENASE; METHANOGENIC ARCHAEBACTERIA; PURIFICATION; CYCLOHYDROLASE; 110-DEGREES-C; HYDROGENASE; REDUCTASE; BACTERIA; COENZYME-F(420); TETRAHYDROMETHANOPTERIN; HYDROGENASE; H-2-FORMING METHYLENETETRAHYDROMETHANOPTERIN DEHYDROGENASE; METHANOBACTERIUM-THERMOAUTOTROPHICUM; METHANOSARCINA-BARKERI; ARCHAEOGLOBUS-FULGIDUS |
| Depositing User: | Dr. Gernot Deinzer |
| Last Modified: | 19 Oct 2022 08:42 |
| URI: | https://pred.uni-regensburg.de/id/eprint/53785 |
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