Purification and subunit characterization of propanediol dehydratase, a membrane-associated enzyme

Abstract

A new isolation procedure for propanediol dehydratase increases by a factor of about 16 the yield of enzyme obtainable from Klebsiella pneumoniae; the enzyme thus isolated has a specific activity of 95 ± 4 units/mg. The apoenzyme consists of four subunits with molecular weights of 60 000, 51 000, 29 000, and 15 000 (±5%). In this new procedure, care was taken to prevent the partial proteolysis of the propanediol dehydratase which seems to occur in earlier procedures. The other novel aspect recognizes that the enzyme is associated with the cell membrane. Accordingly, after gentle sonication, the membrane fragments are separated from cytosol, and the enzyme is solubilized by extraction with buffers containing detergent. The 60 000-dalton subunit has been purified and a partial sequence (34 of the 36 N-terminal residues) determined.

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