Purification and characterization of an L-amino amidase from Mycobacterium neoaurum ATCC 25795

An L-amino amidase from Mycobacterium neoaurum ATCC 25795 responsible for the enantioselective resoln. of DL-a-Me valine amide was purified and characterized. The purifn. procedure included ammonium sulfate fractionation, gel filtration, and anion-exchange chromatog., which resulted in a homogeneous prepn. of the enzyme with a native mol. mass of 136 kDa and a subunit mol. mass of 40 kDa. The purified enzyme displayed the highest activity at 50 DegC and a 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 a-H- and a-alkyl-substituted amino acid amides, with the highest activity towards the cyclo 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-a-allyl alanine amide, DL-a-Me phenylalanine amide, and DL-a-Me leucine amide. [on SciFinder (R)]