Abstract
Methanopyrus kandleri belongs to a novel group of abyssal methanogenic archaebacteria that can grow at 110°C on H2 and CO2 and that shows no close phylogenetic relationship to any methanogen known so far. Methyl-coenzyme M reductase, the enzyme catalyzing the methane forming step in the energy metabolism of methanogens, was purified from this hyperthermophile. The yellow protein with an absorption maximum at 425 nm was found to be similar to the methyl-coenzyme M reductase from other methanogenic bacteria in that it was composed each of two α-, β- and γ-subunits and that it contained the nickel porphinoid coenzyme F430 as prosthetic group. The purified reductase was inactive. The N-terminal amino acid sequence of the γ-subunit was determined. A comparison with the N-terminal sequences of the γ-subunit of methyl-coenzyme M reductases from other methanogenic bacteria revealed a high degree of similarity.
Besides methyl-coenzyme M reductase cell extracts of M. kandleri were shown to contain the following enzyme activities involved in methanogenesis from CO2 (apparent Vmax at 65°C): formylmethanofuran dehydrogenase, 0.3 U/mg protein; formyl-methanofuran: tetrahydromethanopterin formyltransferase, 13 U/mg; N 5,N10-methenyltetrahydromethanopterin cyclohydrolase, 14 U/mg; N 5,N10-methylenetetrahydromethanopterin dehydrogenase (H2-forming), 33 U/mg; N 5,N10-methylenetetrahydromethanopterin reductase (coenzyme F420 dependent), 4 U/mg; heterodisulfide reductase, 2 U/mg; coenzyme F420-reducing hydrogenase, 0.01 U/mg; and methylviologen-reducing hydrogenase, 2.5 U/mg. Apparent Km values for these enzymes and the effect of salts on their activities were determined.
The coenzyme F420 present in M. kandleri was identified as coenzyme F420-2 with 2 γ-glutamyl residues.
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Abbreviations
- H−S-CoM:
-
coenzyme M
- CH3−S-CoM:
-
methylcoenzyme M
- H−S-HTP:
-
7-mercaptoheptanoylthreonine phosphate
- MFR:
-
methanofuran
- CHO-MFR:
-
formyl-MFR
- H4MPT:
-
tetrahydromethanopterin
- CHO−H4MPT:
-
N 5-formyl-H4MPT
- CH=H4MPT+ :
-
N 5,N10-methenyl-H4MPT
- CH2=H4MPT:
-
N 5,N10-methylene-H4MPT
- CH3−H4MPT:
-
N 5-methyl-H4MPT
- F420 :
-
coenzyme F420
- 1 U=:
-
1 μmol/min
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Rospert, S., Breitung, J., Ma, K. et al. Methyl-coenzyme M reductase and other enzymes involved in methanogenesis from CO2 and H2 in the extreme thermophile Methanopyrus kandleri . Arch. Microbiol. 156, 49–55 (1991). https://doi.org/10.1007/BF00418187
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DOI: https://doi.org/10.1007/BF00418187