Characterization of a Novel Thermostable O -Acetylserine Sulfhydrylase from Aeropyrum pernix K1

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Abstract
An O -acetylserine sulfhydrylase (OASS) from the hyperthermophilic archaeon Aeropyrum pernix K1, which shares the pyridoxal 5′-phosphate binding motif with both OASS and cystathionine β-synthase (CBS), was cloned and expressed by using Escherichia coli Rosetta(DE3). The purified protein was a dimer and contained pyridoxal 5′-phosphate. It was shown to be an enzyme with CBS activity as well as OASS activity in vitro. The enzyme retained 90% of its activity after a 6-h incubation at 100°C. In the O -acetyl- l -serine sulfhydrylation reaction, it had a pH optimum of 6.7, apparent K m values for O -acetyl- l -serine and sulfide of 28 and below 0.2 mM, respectively, and a rate constant of 202 s −1 . In the l -cystathionine synthetic reaction, it showed a broad pH optimum in the range of 8.1 to 8.8, apparent K m values for l -serine and l -homocysteine of 8 and 0.51 mM, respectively, and a rate constant of 0.7 s −1 . A. pernix OASS has a high activity in the l -cysteine desulfurization reaction, which produces sulfide and S -(2,3-hydroxy-4-thiobutyl)- l -cysteine from l -cysteine and dithiothreitol.

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