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. 1994 Jan;60(1):153–159. doi: 10.1128/aem.60.1.153-159.1994

Purification and Characterization of an l-Amino Amidase from Mycobacterium neoaurum ATCC 25795

H F M Hermes 1, R F Tandler 1, T Sonke 1, L Dijkhuizen 2,*, E M Meijer 1
PMCID: PMC201283  PMID: 16349147

Abstract

An l-amino amidase from Mycobacterium neoaurum ATCC 25795 responsible for the enantioselective resolution of dl-α-methyl valine amide was purified and characterized. The purification procedure included ammonium sulfate fractionation, gel filtration, and anion-exchange chromatography, which resulted in a homogeneous preparation of the enzyme with a native molecular mass of 136 kDa and a subunit molecular mass of 40 kDa. The purified enzyme displayed the highest activity at 50°C and at pH 8.0 and 9.5. The enzyme was strongly inhibited by the metal-chelating agent 1,10-phenanthroline, the disulfide-reducing agent dithiothreitol, and the cysteine proteinase inhibitor iodoacetamide. The purified amino amidase showed a unique l-enantioselective activity towards a broad range of both α-H- and α-alkyl-substituted amino acid amides, with the highest activity towards the cyclic amino acid amide dl-proline amide. No activity was measured with dl-mandelic acid amide nor with the dipeptide l-phenylalanine-l-leucine. The highest catalytic efficiency (kcat/Km ratio) was measured with dl-α-allyl alanine amide, dl-α-methyl phenylalanine amide, and dl-α-methyl leucine amide.

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Selected References

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