Open AccessS ‐Adenosylmethionine synthetase in the thermophilic archaebacterium Sulfolobus solfataricus .
Author(s) -
PORCELLI Marina,
CACCIAPUOTI Giovanna,
CARTENÌFARINA Maria,
GAMBACORTA Agata
Publication year - 1988
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1988.tb14372.x
Subject(s) - sulfolobus solfataricus , methionine adenosyltransferase , thermophile , biochemistry , divalent , enzyme , isozyme , gene isoform , sulfolobus , methionine , size exclusion chromatography , enzyme kinetics , chemistry , biology , active site , amino acid , archaea , organic chemistry , gene
Two isoforms of methionine adenosyltransferase ( S ‐adenosylmethionine synthetase), A and B, have been partially purified from Sulfolobus solfataricus , a thermophilic archaebacterium optimally growing at 87°C. The chromatographic procedure, involving hydrophobic chromatography on a phenyl‐Sepharose column as a major step. results in 330‐fold and 150‐fold purification of adenosylmethionine synthetase A and B respectively. The apparent molecular masses, estimated by gel filtration, are 180 kDa for A and 75 kDa for B. The A and B isoforms follow Michaelis‐Menten kinetics with apparent K m values of 10 μM nd 20 μM for L‐methionine and of 50 μM and 150 μM for ATP respectively. Adenosylmethionine, a product of the reaction, acts as a powerful non‐competitive inhibitor ( K i = 50 μM) of the A isoform while it inhibits only slightly the B isoform. Both isozymes exhibit tripolyphosphatase activity but only that associated with the form A is stimulated by 5 μM adenosylmethionine concentration. The two enzymes absolutely require a divalent cation for the activity, but are not affected by monovalent ions and reducing agents. The optimum temperature is 90°C and no significant loss of activity is observable after incubation of the two isoforms at 100°C in the presence of ATP. The Arrhenius plots observed for both isozymes are biphasic, indicating different activation energies below and above 75°C. The cytoplasmic levels of ATP, methionine and adenosylmethionine are evaluated.