IP Library Granted Patent US 7,671,189
Granted Patent B2
US 7,671,189 · App. 11/418,828 · Granted Mar 2, 2010

Enzymes having dehalogenase activity and methods of use thereof

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Quick Facts
Patent No.
US 7,671,189
App. No.
11/418,828
Granted
Mar 2, 2010
Kind
B2
Abstract

The invention relates to haloalkane dehalogenases and to polynucleotides encoding the haloalkane dehalogenases. In addition methods of designing new dehalogenases and method of use thereof are also provided. The dehalogenases have increased activity and stability at increased pH and temperature.

Claims (28)

1. An isolated, synthetic or recombinant nucleic acid comprising

(a) a nucleic acid sequence having at least 98% sequence identity to SEQ ID NO:43; or

(b) a sequence completely complementary to (a); or

(c) the nucleic acid of (a), wherein the nucleic acid encodes at least one polypeptide having a dehalogenase activity or encodes at least one polypeptide capable of generating an antibody response by administering the polypeptide to an animal.

2. The isolated, synthetic or recombinant nucleic acid of claim 1 , wherein the nucleic acid sequence has at least 99% sequence identity to SEQ ID NO:43.

3. The isolated, synthetic or recombinant nucleic acid of claim 1 , wherein the nucleic acid sequence has a sequence identity as set forth in SEQ ID NO:43.

4. The isolated, synthetic or recombinant nucleic acid of claim 1 , wherein the nucleic acid sequence encodes a polypeptide having a sequence as set forth in SEQ ID NO:44.

5. The isolated, synthetic or recombinant nucleic acid of claim 1 , wherein the dehalogenase activity comprises catalyzing the conversion of a dihalopropionic acid to an optically active halolactic acid.

6. A vector comprising a nucleic acid as set forth in claim 1 .

7. The vector of claim 6 , wherein the vector is a cloning vector or an expression vector, wherein optionally the expression vector is a mammalian expression vector.

8. The vector of claim 6 , wherein the vector is selected from the group consisting of viral vectors, plasmid vectors, phage vectors, phagemid vectors, cosmids, fosmids, bacteriophages, artificial chromosomes, adenovirus vectors, retroviral vectors, and adeno-associated viral vectors.

9. A host cell comprising a nucleic acid as set forth in claim 1 .

10. The host cell of claim 9 , wherein the host cell is a prokaryote cell, a eukaryote cell, a bacterial cell, a fungal cell, a yeast cell, a mammalian cell, a plant cell or an insect cell.

11. A method for generating a dehalogenase-encoding nucleic acid comprising

(a) providing a parent nucleic acid encoding a dehalogenase comprising a sequence as set forth in claim 1 ; and

(b) modifying the sequence of the parent nucleic acid by a process comprising error-prone PCR, shuffling, oligonucleotide-directed mutagenesis, assembly PCR, sexual PCR mutagenesis, in vivo mutagenesis, cassette mutagenesis, recursive ensemble mutagenesis, exponential ensemble mutagenesis, site-specific mutagenesis, gene reassembly, Gene Site Saturation Mutagenesis (GSSM) or any combination, permutation or iterative process thereof, thereby generating a dehalogenase-encoding nucleic acid.

12. A method for producing a dehalogenase comprising

(a) providing a nucleic acid encoding a dehalogenase comprising a sequence as set forth in claim 1 ; and

(b) expressing the nucleic acid to produce a dehalogenase, wherein optionally the nucleic acid is expressed in a host cell.

13. A method for producing a halolactic acid comprising

(a) providing a nucleic acid encoding a dehalogenase comprising a sequence as set forth in claim 1 , and a dihalopropionic acid;

(b) expressing the nucleic acid to produce a dehalogenase; and

(c) contacting the dihalopropionic acid with the dehalogenase under conditions to produce a halolactic acid, wherein optionally the halolactic acid is an optically active halolactic acid.

14. The nucleic acid of claim 1 bound to a solid matrix or a solid phase.

15. The nucleic acid of claim 14 , wherein the solid phase comprises a glass, a polymeric surface, a packed column of polymeric beads, or a magnetic or a paramagnetic particle.

16. The nucleic acid of claim 1 further comprising an antigen or hapten, an antibody, a specific binding fragment, a biotin, an iminobiotin, an avidin, a streptavidin, a sugar, a lectin, an enzyme, an apoenzyme, a cofactor, a complementary homopolymeric oligonucleotide, a hormone, a receptor.

17. The nucleic acid of claim 1 further comprising a heterologous promoter, wherein optionally the heterologous promoter comprises a eukaryotic promoter, a fungal promoter, a mammalian promoter, a prokaryotic promoter or a viral promoter.

18. A host cell comprising the vector of claim 6 .

Assignments (6)
RELEASE OF SECURITY INTEREST Recorded Jan 9, 2014
From: ATHYRIUM OPPORTUNITIES FUND (A) LP
To: VERENIUM CORPORATION
Reel/Frame 031956/0016 →
NOTICE OF GRANT OF SECURITY INTEREST IN PATENTS Recorded Dec 11, 2012
From: VERENIUM CORPORATION
To: ATHYRIUM OPPORTUNITIES FUND (A) LP
Reel/Frame 029444/0119 →
RELEASE OF SECURITY INTEREST Recorded Jun 1, 2012
From: COMERICA BANK
To: VERENIUM CORPORATION
Reel/Frame 028300/0200 →
SECURITY AGREEMENT Recorded Oct 21, 2011
From: VERENIUM CORPORATION, A DELAWARE CORPORATION
To: COMERICA BANK, A TEXAS BANKING ASSOCIATION
Reel/Frame 027099/0408 →
CHANGE OF NAME Recorded Jan 23, 2008
From: DIVERSA CORPORATION
To: VERENIUM CORPORATION
Reel/Frame 020403/0142 →
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Jan 22, 2008
From: SHORT, JAY M.; RICHARDSON, TOBY; ROBERTSON, DAN E.; GRAY, KEVIN A.
To: DIVERSA CORPORATION
Reel/Frame 020394/0978 →