Open AccessFormylmethanofuran dehydrogenases from methanogenic Archaea Substrate specificity, EPR properties and reversible inactivation by cyanide of the molybdenum or tungsten iron‐sulfur proteins
Author(s) -
BERTRAM Peter A.,
KARRASCH Marion,
SCHMITZ Ruth A.,
BÖCHER Reinhard,
ALBRACHT Simon P. J.,
THAUER Rudolf K.
Publication year - 1994
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1994.tb18646.x
Subject(s) - molybdenum , formate dehydrogenase , chemistry , pterin , electron paramagnetic resonance , cyanide , sulfur , enzyme , cofactor , inorganic chemistry , biochemistry , organic chemistry , nuclear magnetic resonance , physics
Formylmethanofuran dehydrogenases, which are found in methanogenic Archaea, are molybdenum or tungsten iron‐sulfur proteins containing a pterin cofactor. We report here on differences in substrate specificity, EPR properties and susceptibility towards cyanide inactivation of the enzymes from Methanosarcina barkeri, Methanobacterium thermoautotrophicum and Methanobacterium wolfei . The molybdenum enzyme from M. barkeri (relative activity with N ‐formylmethanofuran = 100%) was found to catalyze, albeit at considerably reduced apparent V max , the dehydrogenation of N ‐furfurylformamide (11%), N ‐methylformamide (0.2%), formamide (0.1%) and formate (1%). The molybdenum enzyme from M. Wolfei could only use N ‐furfurylformamide (1%) and formate (3%) as pseudosubstrates. The molybdenum enzyme from M. thermoautotrophicum and the tungsten enzymes from M. thermoautotrophicum and M. wolfei were specific for N ‐formylmethanofuran. The molybdenum formylmethanofuran dehydrogenases exhibited at 77 K two rhombic EPR signals, designated FMD red and FMD ox , both derived from Mo as shown by isotopic substitution with 97 Mo. The FMD red signal was only displayed by the active enzyme in the reduced form and was lost upon enzyme oxidation; the FMD ox signal was displayed by an inactive form and was not quenched by O 2 . The tungsten isoenzymes were EPR silent. The molybdenum formylmethanofuran dehydrogenases were found to be inactivated by cyanide whereas the tungsten isoenzymes, under the same conditions, were not inactivated. Inactivation was associated with a characteristic change in the molybdenum‐derived EPR signal. Reactivation was possible in the presence of sulfide.