Open AccessCharacterization of a new cobalt‐containing nitrile hydratase purified from urea‐induced cells of Rhodococcus rhodochrous J1
Author(s) -
NAGASAWA Toru,
TAKEUCHI Koji,
YAMADA Hideaki
Publication year - 1991
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1991.tb15853.x
Subject(s) - nitrile hydratase , rhodococcus rhodochrous , nitrile , chemistry , enzyme , nitrilase , rhodococcus , urea , ferric , protein subunit , biochemistry , stereochemistry , organic chemistry , gene
A new cobalt‐containing nitrile hydratase was purified from extracts of urea‐induced cells from Rhodococcus rhodochrous J1 in seven steps. At the last step, the enzyme was crystallized by adding ammonium sulfate. Nitrile hydratase was a 500–530‐kDa protein composed of two different subunits (α subunit 26kDa, β subunit 29kDa). The enzyme contained approximately 11–12 mol cobalt/mol enzyme. A concentrated solution of highly purified nitrile hydratase exhibited a broad absorption spectrum in the visible range, with an absorption maxima at 410 nm. The enzyme had a wide substrate specificity. Aliphatic saturated or unsaturated nitriles as well as aromatic nitriles, were substrates for the enzyme. The optimum pH of the hydratase was pH 6.5–6.8. The enzyme was more stable than ferric nitrile hydratases. The amino‐terminal sequence of each subunit of R. rhodochrous J1 enzyme was determined and compared with that of ferric nitrile hydratases. Prominent similarities were observed with the β subunit. However, the amino acid sequence of the α subunit from R. rhodochrous J1 was quite different from that of the ferric enzymes.