O-acetylserine sulfhydrylase from Methanosarcina thermophila

Birthe Borup, James G. Ferry

Research output: Contribution to journalArticle

25 Scopus citations

Abstract

Cysteine is the major source of fixed sulfur for the synthesis of sulfur, containing compounds in organisms of the Bacteria and Eucarya domains. Though pathways for cysteine biosynthesis have been established for both of these domains, it is unknown how the Archaea fix sulfur or synthesize cysteine. None of the four archaeal genomes sequenced to date contain open reading frames with identities to either O-acetyl-L-serine sulfhydrylase (OASS) or homocysteine synthase, the only sulfur-fixing enzymes known in nature. We report the purification and characterization of OASS from acetate- grown Methanosarcina thermophila, a moderately thermophilic methanoarchaeon. The purified OASS contained pyridoxal 5'-phosphate and catalyzed the formation of L-cysteine and acetate from O-acetyl-L-serine and sulfide. The N-terminal amino acid sequence has high sequence similarity with other known OASS enzymes from the Eucarya and Bacteria domains. The purified OASS had a specific activity of 129 μmol of cysteine/min/mg, with a K(m) of 500 ± 80 μM for sulfide, and exhibited positive cooperativity and substrate inhibition with O-acetyl-L-serine. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a single band at 36 kDa, and native gel filtration chromatography indicated a molecular mass of 93 kDa, suggesting that the purified OASS is either a homodimer or a homotrimer. The optimum temperature for activity was between 40 and 60°C, consistent with the optimum growth temperature for M. thermophila. The results of this study provide the first evidence for a sulfur-fixing enzyme in the Archaea domain. The results also provide the first biochemical evidence for an enzyme with the potential for involvement in cysteine biosynthesis in the Archaea.

Original languageEnglish (US)
Pages (from-to)45-50
Number of pages6
JournalJournal of bacteriology
Volume182
Issue number1
DOIs
StatePublished - Jan 2000

All Science Journal Classification (ASJC) codes

  • Microbiology
  • Molecular Biology

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