US 2003/0157677 A1 · Mutants of Mycobacterium vaccae-derived formate dehydrogenase and uses thereof

Abstract

An objective of the present invention is to provide polypeptides capable of retaining a strong enzyme activity of formate dehydrogenase in the presence of an organic solvent and to provide the uses thereof. Formate dehydrogenase mutant polypeptides, which are resistant to organic solvents, were constructed by substituting cysteines at position 146 and/or at position 256 in the amino acid sequence of Mycobacterium vaccae-derived formate dehydrogenase by site-directed mutagenesis. The polypeptides have strong activities of formate dehydrogenase in the presence of an organic solvent. The mutants are useful for the production of alcohols using ketones as raw material, etc.

Claims29

Dependent claims

An isolated polypeptide having a strong formate dehydrogenase activity in the presence of an organic solvent, said polypeptide comprising a mutation in which the cysteine residue at position 146 or position 256 in the amino acid sequence of SEQ ID NO:2 is substituted with an amino acid other than cysteine.

An isolated polypeptide selected from the group consisting of:

(1) the sequence of SEQ ID NO:2 in which cysteines at positions 6, 146, and 256 have been substituted with serine;
(2) the sequence of SEQ ID NO:2 in which cysteine at position 6 has been substituted with alanine, and cysteine at position 256 has been substituted with serine;
(3) the sequence of SEQ ID NO:2 in which cysteine at position 6 has been substituted with valine, and cysteine at position 256 has been substituted with serine;
(4) the sequence of SEQ ID NO:2 in which cysteine at position 6 has been substituted with serine, and cysteine at position 256 has been substituted with alanine;
(5) the sequence of SEQ ID NO:2 in which cysteine at position 6 has been substituted with serine, and cysteine at position 256 has been substituted with valine;
(6) the sequence of SEQ ID NO:2 in which cysteine at position 146 has been substituted with serine;
(7) the sequence of SEQ ID NO:2 in which cysteine at position 256 has been substituted with serine;
(8) the sequence of SEQ ID NO:2 in which cysteines at positions 146 and 256 have been substituted with serine;
(9) the sequence of SEQ ID NO:2 in which cysteine at position 256 has been substituted with valine;
(10) the sequence of SEQ ID NO:2 in which cysteine at position 146 has been substituted with serine, and cysteine at position 256 has been substituted with valine;
(11) the sequence of SEQ ID NO:2 in which cysteine at position 6 has been substituted with alanine, and cysteine at position 256 has been substituted with valine;
(12) the sequence of SEQ ID NO:2 in which cysteine at position 6 has been substituted with alanine, cysteine at position 146 has been substituted with serine, and cysteine at position 256 has been substituted with valine;
and
(13) the sequence of SEQ ID NO:2 in which cysteines at positions 6 and 146 have been substituted with alanines, and cysteine at position 256 has been substituted with valine.

11 more elements

Dependencies are inferred from the claim text.

Mutants of Mycobacterium vaccae-derived formate dehydrogenase and uses thereof

Life of this patent

Mutants of Mycobacterium vaccae-derived formate dehydrogenase and uses thereof

Life of this patent

Abstract

Claims29