Summary
Production, purification and characterization of the NAD(H)-dependent d-mandelate dehydrogenase from Lactobacillus curvatus was studied. An enzyme level of about 150 U/1 could be obtained by anaerobic cultivation in liquid broth. The specific enzyme activity in the crude extract was 1—3 U/mg. Purification by liquidliquid extraction and ion exchange chromatography led to a preparation of 2100 U/mg. The molecular weight of the enzyme was determined to be 60000 (gel filtration on Superose S12) containing two subunits of 30000. A variety of aliphatic and aromatic α-keto acids are accepted as substrates by the mandelate dehydrogenase, for the substrate benzoylformate a Michaelis constant of 2·10-4M was measured. Cu2+-ions and mercury compounds such as HgCl2 or p-chloromercuribenzoate are strong inhibitors at concentrations of 0.1 mM. An unoptimized continuous conversion in an enzyme-membrane-reactor demonstrated that the enzyme could be applied for the stereospecific synthesis of d-mandelic acid.
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Abbreviations
- FDH:
-
formate dehydrogenase
- PEG:
-
polyethylenglycol
- SDS:
-
sodiumdodecylsulfate
- MRS:
-
growth medium according to deMan, Rogosa and Sharpe (deMan et al. 1960)
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Hummel, W., Schütte, H. & Kula, MR. d-(-)-Mandelic acid dehydrogenase from Lactobacillus curvatus . Appl Microbiol Biotechnol 28, 433–439 (1988). https://doi.org/10.1007/BF00268209
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DOI: https://doi.org/10.1007/BF00268209