Open AccessA novel nitrilase, arylacetonitrilase, of Alcaligenes faecalis JM3
Author(s) -
NAGASAWA Toru,
MAUGER Jacques,
YAMADA Hideaki
Publication year - 1990
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1990.tb19467.x
Subject(s) - nitrilase , alcaligenes faecalis , chemistry , molecular mass , enzyme , dithiothreitol , alcaligenes , hydrolysis , amidase , nitrile hydratase , chromatography , agarose , polyacrylamide gel electrophoresis , gel electrophoresis , nitrile , amide , biochemistry , organic chemistry , bacteria , pseudomonas , biology , genetics
A new type of nitrilase, arylacetonitrilase, has been purified from isovaleronitrile‐induced cells of Alcaligenes faecalis JM3 in four steps. The purity of the enzyme was confirmed by SDS/polyacrylamide gel electrophoresis, ampholyte electrofocusing and double immunodiffusion in agarose. The enzyme has a molecular mass of about 275 kDa and consists of six subunits of identical molecular mass. The purified enzyme exhibits a pH optimum of 7.5 and a temperature optimum of 45°C. The enzyme is specific for arylacetonitriles such as 2‐thiophenacetonitrile, p ‐tolylacetonitrile, p ‐chlorobenzylcyanide, p ‐fluorobenzylcyanide and 3‐pyridylacetonitrile. The enzyme stoichio‐metrically catalyzes the hydrolysis of arylacetonitrile to arylacetic acid and ammonia, no formation of amide occurring. However, the enzyme does not attack nitrile groups attached to aromatic and heteroaromatic rings, which are hydrolyzed preferably by the nitrilases known previously. The enzyme requires thiol compounds such as dithiothreitol and 2‐mercaptoethanol to exhibit its maximum activity.