Functional Expression of Nitrile Hydratase in Escherichia coli: Requirement of a Nitrile Hydratase Activator and Post-Translational Modification of a Ligand Cysteine.

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概要

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• The nitrile hydratase (NHase) from Rhodococcus sp. N-771 is a photoreactive enzyme that is inactivated on nitrosylation of the non-heme iron center and activated on photo-dissociation of nitric oxide (NO). The nitrite hydratase operon consists of six genes encoding NHase regulator 2, NHase regulator 1, amidase, NHase α subunit, NHase β subunit and NHase activator. We overproduced the NHase in Escherichia coli using a T 7 expression system. The NHase was functionally expressed in E. coli only when the NHase activator encoded downstream of the β subunit gene was co-expressed and the transformant was grown at 30°C or less. A ligand cysteine, αCys 112, of the recombinant NHase was also posttranslationally modified to a cysteine-sulfinic acid similar to for the native NHase. Although another modification of αCys 114 could not be identified because of the instability under acidic conditions, the recombinant NHase could be reversibly inactivated by nitric oxide.

• 社団法人 日本生化学会の論文

著者

• Nojiri Masaki

  The Institute Of Physical And Chemial Research (riken)

• Takio Koji

  The Institute of physical and Chemical Research(RIKEN)

• YOHDA Masafumi

  The Institute of Physical and Chemial Research (RIKEN)

• ENDO Isao

  The Institute of Physical and Chemical Research (RIKEN)

• Odaka Masafumi

  The Institute of Physical and Chemial Research (RIKEN)

• Dohmae Naoshi

  The Institute of Physical and Chemical Research (RIKEN)

• Matsushita Yusuke

  The Institute of Physical and Chemical Research (RIKEN)

• Tsujimura Masanari

  The Institute of Physical and Chemical Research (RIKEN)

• Yoshida Takao

  The Institute of Physical and Chemical Research (RIKEN)

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• Functional Expression of Nitrile Hydratase in Escherichia coli: Requirement of a Nitrile Hydratase Activator and Post-Translational Modification of a Ligand Cysteine.

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