Chemistry: molecular biology and microbiology – Enzyme – proenzyme; compositions thereof; process for... – Oxidoreductase
Reexamination Certificate
1999-05-18
2001-06-19
Slobodyansky, Elizabeth (Department: 1652)
Chemistry: molecular biology and microbiology
Enzyme , proenzyme; compositions thereof; process for...
Oxidoreductase
C435S191000, C435S252300, C435S320100, C536S023200
Reexamination Certificate
active
06248575
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates to isolated nucleic acid sequences encoding polypeptides having L-amino acid oxidase activity. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the nucleic acid sequences as well as recombinant methods for producing the polypeptides.
2. Description of the Related Art
Polypeptides having L-amino acid oxidase activity catalyze the oxidative deamination of L-amino acids to produce the corresponding 2-keto acids. Such polypeptides are classified under the Enzyme Classification Number E.C. 1.4.3.2 of the International Union of Biochemistry and Molecular Biology.
WO 94/25574 discloses an L-amino acid oxidase from
Trichoderma harzianum
which has a broad substrate specificity. JP 78115867A discloses an L-amino acid oxidase from
Trichoderma viride
which has a very high substrate specificity toward L-lysine.
Niedermann and Lerch (1990,
Journal of Biological Chemistry
265: 17246-17251) disclose the molecular cloning of the L-amino acid oxidase gene from
Neurospora crassa.
Bockholt et al. (1995,
Biochimica et Biophysica Acta
1264: 289-293) disclose the cloning and DNA sequence analysis of the L-amino acid oxidase gene from the cyanobacterium Synechococcus PCC6301.
It is an object of the present invention to provide new isolated nucleic acid sequences encoding polypeptides having L-amino acid oxidase activity.
SUMMARY OF THE INVENTION
The present invention relates to isolated nucleic acid sequences encoding polypeptides having L-amino acid oxidase activity, selected from the group consisting of:
(a) a nucleic acid sequence encoding a polypeptide having an amino acid sequence which has at least 50% identity with amino acids 1 to 617 of SEQ ID NO. 2;
(b) a nucleic acid sequence having at least 50% homology with nucleotides 1 to 1851 of SEQ ID NO. 1;
(c) a nucleic acid sequence which hybridizes under low stringency conditions with (i) nucleotides 1 to 1851 of SEQ ID NO. 1, (ii) the genomic DNA sequence comprising nucleotides 1 to 1851 of SEQ ID NO. 1, (iii) a subsequence of (i) or (ii) of at least about 100 nucleotides, or (iv) a complementary strand of (i), (ii), or (iii);
(d) a nucleic acid sequence encoding a variant of the polypeptide having an amino acid sequence of SEQ ID NO. 2 comprising a substitution, deletion, and/or insertion of one or more amino acids;
(e) an allelic variant of (a), (b), or (c);
(f) a subsequence of (a), (b), (c), or (e), wherein the subsequence encodes a polypeptide fragment which has L-amino acid oxidase activity; and
(g) a nucleic acid sequence encoding a polypeptide having L-amino acid oxidase activity with physicochemical properties of (i) a pH optimum in the range of from about pH 8.5 to about pH 9.5 determined after incubation for 20 minutes at 20° C. in the presence of L-arginine; (ii) a pH stability of 80% or more, relative to initial activity, at pH 9.5 determined after incubation for 1 hour at 40° C. in the absence of substrate; and (iii) an activity towards L-arginine, L-lysine, L-methionine, L-asparagine, L-phenylalanine, and/or L-leucine.
The present invention also relates to nucleic acid constructs, vectors, and host cells comprising the nucleic acid sequences as well as recombinant methods for producing the polypeptides.
REFERENCES:
patent: 5536655 (1996-07-01), Thomas et al.
patent: 5801035 (1998-09-01), Schneider et al.
patent: 5817499 (1998-10-01), Dalboge et al.
patent: 55-43409 (1990-03-01), None
patent: 94/25574 (1994-11-01), None
Niedermann and Lerch, 1990, Journal of biological Chemistry 265: 17246-17251.
Bockholt et al., 1995, Biochimica et Biophysica Acta 1264: 289-293.
Novozymes Biotech Inc.
Slobodyansky Elizabeth
Starnes Robert L.
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