Granted Patent
B2
US 12,286,648 · App. 18/180,711 · Granted Apr 29, 2025
Engineered proline hydroxylase biocatalysts for hydroxylation of chemical compounds
Inventors:
Jovana Nazor (Milpitas, CA); Robert Osborne (Raleigh, NC); Jack Liang (South San Francisco, CA); Jonathan Vroom (South San Francisco, CA); Xiyun Zhang (Fremont, CA); David Entwistle (San Carlos, CA); Rama Voladri (Pleasanton, CA); Ravi David Garcia (Los Gatos, CA); Jeffrey C. Moore (Westfield, NJ); Shane Grosser (Princeton, NJ); Birgit Kosjek (Westfield, NJ); Matthew Truppo (Ocean Township, NJ)
Assignee:
Codexis, Inc.
View Patent ↗
Loading inventors, assignments & file history…
Abstract
The present invention provides engineered proline hydroxylase polypeptides for the production of hydroxylated compounds, polynucleotides encoding the engineered proline hydroxylases, host cells capable of expressing the engineered proline hydroxylases, and methods of using the engineered proline hydroxylases to prepare compounds useful in the production of active pharmaceutical agents.
Claims (16)
1. An engineered polypeptide having proline hydroxylase activity comprises an amino acid sequence having at least 90% sequence identity to reference sequence SEQ ID NO:604 and one or more residue differences as compared to SEQ ID NO:604 at residue positions selected from: 13, 14, 24, 26, 27, 30, 57, 61, 62, 72, 76, 77, 81, 82, 86, 88, 97, 114, 127, 128, 142, 158, 161, 163, 173, 175, 176, 178, 180, 184, 185, 186, 187, 188, 189, 191, 192, 195, 198, 200, 207, 209, 210, 211, 213, 215, 217, 218, 222, 225, 230, 233, 236, 238, 240, 241, 256, 259, 263, 265, 271, and 273.
2. The engineered polypeptide of claim 1 , wherein said polypeptide has at least 90% sequence identity to SEQ ID NO: 604 or 640.
3. The engineered polypeptide of claim 1 , wherein said polypeptide comprises residue differences at positions 26, 30, 62, 82, 114, 158, 161, 271, and 273, as compared to SEQ ID NO: 604.
4. The engineered polypeptide of claim 1 , wherein said polypeptide comprises residue differences of 26A, 30N, 62D, 82K, 114S, 158N, 161P, 271W, and 273T, as compared to SEQ ID NO: 604.
5. The engineered polypeptide of claim 1 , wherein said engineered polypeptide is capable of converting(S)-pipecolic acid to (25,55)-5-hydroxypipecolic acid.
6. The engineered polypeptide of claim 5 , wherein said engineered polypeptide is capable of converting(S)-pipecolic acid to (25,55)-5-hydroxypipecolic acid with at least 1.2 fold, 1.5 fold, 2 fold, 3 fold, 4 fold, 5 fold, 10 fold, 20 fold, 30 fold, 40 fold, 50 fold, 60 fold, 70 fold, 80 fold, 90 fold, 100 fold or more the activity of the naturally occurring enzyme.
7. The engineered polypeptide of claim 5 , wherein said engineered polypeptide is capable of converting(S)-pipecolic acid to (2S,5S)-5-hydroxypipecolic acid with greater than 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more diastereomeric excess of (2S,5R)-5-hydroxypipecolic acid.
8. A polynucleotide encoding the engineered polypeptide of claim 1 .
9. A polynucleotide encoding the engineered polypeptide of claim 3 .
10. The polynucleotide of claim 8 , wherein said polynucleotide comprises a nucleic acid sequence optimized for expression in E. coli.
11. The polynucleotide of claim 9 , wherein said polynucleotide comprises a nucleic acid sequence optimized for expression in E. coli.
12. An expression vector comprising the polynucleotide of claim 8 , optionally further comprising at least one control sequence.
13. The expression vector of claim 12 , wherein said vector comprises SEQ ID NO:1007, 1008, or 1009.
14. A host cell comprising the polynucleotide of claim 8 .
15. A method of preparing an engineered polypeptide, comprising culturing the host cell of claim 14 , under conditions suitable for expression of the polypeptide.
16. The method of claim 15 , further comprising a step of isolating the engineered polypeptide.
Assignments (1)
SECURITY INTEREST
Recorded Feb 15, 2024
From: CODEXIS, INC.
To: INNOVATUS LIFE SCIENCES LENDING FUND I, LP, AS COLLATERAL AGENT
Reel/Frame 066600/0650 →
Continuity (5)
Provisional Application
62347724
· Jun 9, 2016
References Cited (244)
US 3768871A
· Meyers
· 1973
[cited by applicant]
US 5364775A
· Katsumata et al.
· 1994
[cited by applicant]
US 5605793A
· Stemmer
· 1997
[cited by applicant]
US 5811238A
· Stemmer et al.
· 1998
[cited by applicant]
US 5830721A
· Stemmer et al.
· 1998
[cited by applicant]
US 5834252A
· Stemmer et al.
· 1998
[cited by applicant]
US 5837458A
· Minshull et al.
· 1998
[cited by applicant]
US 5854040A
· Ozaki et al.
· 1998
[cited by applicant]
US 5928905A
· Stemmer et al.
· 1999
[cited by applicant]
US 5962292A
· Ozaki et al.
· 1999
[cited by applicant]
US 5963254A
· Kim et al.
· 1999
[cited by applicant]
US 6096548A
· Stemmer
· 2000
[cited by applicant]
US 6117679A
· Stemmer
· 2000
[cited by applicant]
US 6132970A
· Stemmer
· 2000
[cited by applicant]
US 6165793A
· Stemmer
· 2000
[cited by applicant]
US 6180406B1
· Stemmer
· 2001
[cited by applicant]
US 6251674B1
· Tobin et al.
· 2001
[cited by applicant]
US 6277638B1
· Stemmer
· 2001
[cited by applicant]
US 6287861B1
· Stemmer et al.
· 2001
[cited by applicant]
US 6287862B1
· delCardayre et al.
· 2001
[cited by applicant]
US 6291242B1
· Stemmer
· 2001
[cited by applicant]
US 6297053B1
· Stemmer
· 2001
[cited by applicant]
US 6303344B1
· Patten et al.
· 2001
[cited by applicant]
US 6309883B1
· Minshull et al.
· 2001
[cited by applicant]
US 6319713B1
· Patten et al.
· 2001
[cited by applicant]
US 6319714B1
· Crameri et al.
· 2001
[cited by applicant]
US 6323030B1
· Stemmer
· 2001
[cited by applicant]
US 6326204B1
· delCardayre et al.
· 2001
[cited by applicant]
US 6335160B1
· Patten et al.
· 2002
[cited by applicant]
US 6335198B1
· delCardayre et al.
· 2002
[cited by applicant]
US 6344356B1
· Stemmer
· 2002
[cited by applicant]
US 6352859B1
· delCardayre et al.
· 2002
[cited by applicant]
US 6355484B1
· Patten et al.
· 2002
[cited by applicant]
US 6358740B1
· Patten et al.
· 2002
[cited by applicant]
US 6358742B1
· Stemmer
· 2002
[cited by applicant]
US 6365377B1
· Patten et al.
· 2002
[cited by applicant]
US 6365408B1
· Stemmer
· 2002
[cited by applicant]
US 6368861B1
· Crameri et al.
· 2002
[cited by applicant]
US 6372497B1
· Stemmer
· 2002
[cited by applicant]
US 6376246B1
· Crameri et al.
· 2002
[cited by applicant]
US 6379964B1
· delCardayre et al.
· 2002
[cited by applicant]
US 6387702B1
· Stemmer
· 2002
[cited by applicant]
US 6391552B2
· Stemmer
· 2002
[cited by applicant]
US 6391640B1
· Minshull et al.
· 2002
[cited by applicant]
US 6395547B1
· Stemmer
· 2002
[cited by applicant]
US 6406855B1
· Patten et al.
· 2002
[cited by applicant]
US 6406910B1
· Patten et al.
· 2002
[cited by applicant]
US 6413745B1
· Patten et al.
· 2002
[cited by applicant]
US 6413774B1
· Stemmer
· 2002
[cited by applicant]
US 6420175B1
· Stemmer
· 2002
[cited by applicant]
US 6423542B1
· Crameri et al.
· 2002
[cited by applicant]
US 6426224B1
· Crameri et al.
· 2002
[cited by applicant]
US 6436675B1
· Welch et al.
· 2002
[cited by applicant]
US 6444468B1
· Stemmer et al.
· 2002
[cited by applicant]
US 6455253B1
· Patten et al.
· 2002
[cited by applicant]
US 6479652B1
· Crameri et al.
· 2002
[cited by applicant]
US 6482647B1
· Stemmer
· 2002
[cited by applicant]
US 6489146B2
· Stemmer et al.
· 2002
[cited by applicant]
US 6506602B1
· Stemmer
· 2003
[cited by applicant]
US 6506603B1
· Stemmer
· 2003
[cited by applicant]
US 6519065B1
· Colbourne et al.
· 2003
[cited by applicant]
US 6521453B1
· Crameri et al.
· 2003
[cited by applicant]
US 6528311B1
· delCardayre et al.
· 2003
[cited by applicant]
US 6537746B2
· Amold et al.
· 2003
[cited by applicant]
US 6573098B1
· Stemmer
· 2003
[cited by applicant]
US 6576467B1
· Stemmer
· 2003
[cited by applicant]
US 6579678B1
· Patten et al.
· 2003
[cited by applicant]
US 6586182B1
· Patten et al.
· 2003
[cited by applicant]
US 6602986B1
· Stemmer et al.
· 2003
[cited by applicant]
US 6613514B2
· Patten et al.
· 2003
[cited by applicant]
US 6653072B1
· Patten et al.
· 2003
[cited by applicant]
US 6716631B1
· delCardayre et al.
· 2004
[cited by applicant]
US 6946296B2
· Patten et al.
· 2005
[cited by applicant]
US 6961664B2
· Selifonov et al.
· 2005
[cited by applicant]
US 6995017B1
· Stemmer
· 2006
[cited by applicant]
US 7024312B1
· Selifonov et al.
· 2006
[cited by applicant]
US 7058515B1
· Selifonov et al.
· 2006
[cited by applicant]
US 7105297B2
· Minshull et al.
· 2006
[cited by applicant]
US 7148054B2
· delCardayre et al.
· 2006
[cited by applicant]
US 7288375B2
· Stemmer et al.
· 2007
[cited by applicant]
US 7421347B2
· Selifonov et al.
· 2008
[cited by applicant]
US 7430477B2
· Selifonov et al.
· 2008
[cited by applicant]
US 7534564B2
· Patten et al.
· 2009
[cited by applicant]
US 7620500B2
· Mundorff et al.
· 2009
[cited by applicant]
US 7620502B2
· Selifonov et al.
· 2009
[cited by applicant]
US 7629170B2
· delCardayre et al.
· 2009
[cited by applicant]
US 7702464B1
· Emig et al.
· 2010
[cited by applicant]
US 7747391B2
· Gustafsson et al.
· 2010
[cited by applicant]
US 7747393B2
· Fox
· 2010
[cited by applicant]
US 7751986B2
· Gustafsson et al.
· 2010
[cited by applicant]
US 7776598B2
· Patten et al.
· 2010
[cited by applicant]
US 7783428B2
· Gustafsson et al.
· 2010
[cited by applicant]
US 7795030B2
· Minshull et al.
· 2010
[cited by applicant]
US 7853410B2
· Selifonov et al.
· 2010
[cited by applicant]
US 7868138B2
· Stemmer et al.
· 2011
[cited by applicant]
US 7873499B2
· Selifonov et al.
· 2011
[cited by applicant]
US 7904249B2
· Selifonov et al.
· 2011
[cited by applicant]
US 7957912B2
· Selifonov et al.
· 2011
[cited by applicant]
US 8383346B2
· Colbeck et al.
· 2013
[cited by applicant]
US 8504498B2
· Fox
· 2013
[cited by applicant]
US 8762066B2
· Fox
· 2014
[cited by applicant]
US 8849575B2
· Gustafsson et al.
· 2014
[cited by applicant]
US 9790527B2
· Chen et al.
· 2017
[cited by applicant]
US 20060195947A1
· Davis et al.
· 2006
[cited by applicant]
US 20080220990A1
· Fox
· 2008
[cited by applicant]
US 20090312196A1
· Colbeck et al.
· 2009
[cited by applicant]
US 20110046102A1
· Ledoussal et al.
· 2011
[cited by applicant]
US 20110091942A1
· Kino et al.
· 2011
[cited by applicant]
CN 104428412A
· 2015
[cited by applicant]
CN 105177026A
· 2015
[cited by applicant]
EP 0641862B1
· 2001
[cited by applicant]
EP 2290065B1
· 2014
[cited by applicant]
EP 3536780A1
· 2019
[cited by applicant]
JP 2014236713A
· 2014
[cited by applicant]
WO 9522625A1
· 1995
[cited by applicant]
WO 9533836A1
· 1995
[cited by applicant]
WO 9600787A1
· 1996
[cited by applicant]
WO 970078A1
· 1997
[cited by applicant]
WO 9735966A1
· 1997
[cited by applicant]
WO 9827230A1
· 1998
[cited by applicant]
WO 200042651A1
· 2000
[cited by applicant]
WO 200175767A2
· 2001
[cited by applicant]
WO 2009008908A2
· 2009
[cited by applicant]
WO 2009091856A2
· 2009
[cited by applicant]
WO 2009102899A1
· 2009
[cited by applicant]
WO 2009102901A1
· 2009
[cited by applicant]
WO 2009139365A1
· 2009
[cited by applicant]
WO 2009152336A1
· 2009
[cited by applicant]
WO 2010126820A2
· 2010
[cited by applicant]
WO 2011035105A1
· 2011
[cited by applicant]
WO 2013003290A1
· 2013
[cited by applicant]
WO 2013138339A1
· 2013
[cited by applicant]
WO 2013159055A1
· 2013
[cited by applicant]
WO 2013169725A2
· 2013
[cited by applicant]
WO 2014120819A1
· 2014
[cited by applicant]
WO 2014120821A1
· 2014
[cited by applicant]
WO 2015048573A1
· 2015
[cited by applicant]
WO 2015098774A1
· 2015
[cited by applicant]
Chica et al. Curr Opin Biotechnol. Aug. 2005;16(4):378-84. (Year: 2005).
[cited by examiner]
Singh et al. Curr Protein Pept Sci. 2017, 18, 1-11 (Year: 2017).
[cited by examiner]
Adams, D.R., et al., “An efficient route to the alpha-methyl ester of L-glutamic acid, and its conversion into cis-5-hydroxy-L-pipecolic acid,” Chem. Commun., 3:349-350 [1996].
[cited by applicant]
Altamura, M., et al., “2-Substituted penems with amino acid-related side chains: synthesis and antibacterial activity of a new series of beta-lactam antibiotics,” J Med Chem., 38(21):4244-56 [1995].
[cited by applicant]
Altschul, S., et al., “Basic local alignment search tool,” J. Mol. Biol., 215: 403-410 (1990).
[cited by applicant]
Altschul, S.F., et al., “Gapped BLAST and PSI-BLAST: a new generation of protein database search programs,” Nucleic Acids Res., 25(17):3389-3402 (1997).
[cited by applicant]
Baldino, Jr., F., et al., “High-Resolution in Situ Hybridization Histochemistry,” Methods Enzymology, 168:761-777 (1989).
[cited by applicant]
Batzer, M.A., “Erratum: Structure and variability of recently inserted Alu family members”, Nucleic Acids Res 19:698-699 [1991].
[cited by applicant]
Beaucage, S.L., et al., “Deoxynucleoside phosphoamidites-A new class of key intermediates for deoxypolynucleotide synthesis,” Tetrahedron Letters, 22(20):1859-62 (1981).
[cited by applicant]
Beyerman, H.C., et al., “Stereospecific synthesis and optical resolution of 5-hydroxypipecolic acid,” Recueil des Travaux Chimiques des Pays-Bas, 78(9):648-658 [1959].
[cited by applicant]
Black, M.E., et al., “Creation of drug-specific herpes simplex virus type 1 thymidine kinase mutants for gene therapy,” Proc Natl Acad Sci USA, 93:3525-3529 (1996).
[cited by applicant]
Bolton, E.T., et al., “A General Method for the lisolation of RNA Complementary to DNA,” Proc. Natl. Acad. Sci. USA 48:1390 ( 1962).
[cited by applicant]
Botman, P.N., et al., “Diastereoselective synthesis of (2S,5R)-5-hydroxypipecolic acid and 6-substituted derivatives,” Organic Letters, 6(26):4941-4944 [2004].
[cited by applicant]
Botstein, D., et al., “Strategies and applications of in vitro mutagenesis,” Science, 229(4719):1193-1201, 1985.
[cited by applicant]
Breslauer, K.J., et al., “Predicting DNA duplex stability from the base sequence,” Proc. Natl. Acad. Sci. USA, 83:3746-3750 (1986).
[cited by applicant]
Caldwell, R.C., et al., “Mutagenic PCR,” PCR Methods Appl., 3:S136-S140 (1994).
[cited by applicant]
Callens, R.E.A., et al., “Preparation of Trans-5-Hydroxy-L-Pipecolic Acid and Cis-4-Hydroxy-L-Pipecolic Acid From L-Baikiain (1,2,5,6-L-Tetrahydropyridine-2-Carboxylic Acid),” Bulletin des Sociétés Chimiques Belges, 91(…
[cited by applicant]
Carter, P., “Site-directed mutagenesis,” Biochem. J., 237:1-7 (1986).
[cited by applicant]
Chen, K.X., et al., “Novel potent hepatitis C virus NS3 serine protease inhibitors derived from proline-based macrocycles,” J Med Chem., 49(3):995-1005 [2006].
[cited by applicant]
Chen, K.X., et al., “Syntheses of novel 4-tert-alkyl ether proline-based 16- and 17-membered macrocyclic compounds,” J Org Chem., 67(8):2730-3 [2002].
[cited by applicant]
Chiou, W.H., et al., “Facile syntheses of enantiopure 3-hydroxypiperidine derivatives and 3-hydroxypipecolic acids,” J Org Chem., 75(5):1748-51 [2010].
[cited by applicant]
Christians, F.C., et al., “Directed evolution of thymidine kinase for AZT phosphorylation using DNA family shuffling,” Nat. Biotechnol., 17:259-264 (1999).
[cited by applicant]
Clark-Lewis, J.W., et al., “Occurrence of 4-hydroxypipecolic acid in Acacia species,” Nature, 184(Suppl 16):1234-5 [1959].
[cited by applicant]
Cohen, L.A., et al., “Synthesis of 5-Hydroxypipecolic Acid and Separation of Its Diastereoisomers,” Science, 123 3202):842-843 [1956].
[cited by applicant]
Crameri, A., et al., “DNA shuffling of a family of genes from diverse species accelerates directed evolution”, Nature, 391:288-291 (1998).
[cited by applicant]
Crameri, A., et al., “Improved green fluorescent protein by molecular evolution using DNA shuffling. ”Nat. Biotechnol., 14(3):315-319 (1996).
[cited by applicant]
Crameri, A., et al., “Molecular evolution of an arsenate detoxification pathway by DNA shuffling,” Nat. Biotechnol., 15(5):436-438 (1997).
[cited by applicant]
Dale, S.J., et al., “Oligonucleotide-directed random mutagenesis using the phosphorothioate method,” Methods Mol. Biol., 57:369-74 (1996).
[cited by applicant]
De Boer, H.A., et al., “The tac promoter: A functional hybrid derived from the trp and lac promoters,” Proc. Natl Acad. Sci. USA, 80: 21-25 (1983).
[cited by applicant]
Efimov, V.A., et al., “Hydroxyproline-based DNA mimics provide an efficient gene silencing in vitro and in vivo,” Nucleic Acids Res., 34(8):2247-2257 [2006].
[cited by applicant]
Eguchi, C., et al., “The novel synthesis of L-Hydroxyproline from D-Glutamic Acid,” Bull. Chem. Soc. Japan, 47(7):1704-08 [1974].
[cited by applicant]
Fasman, G.D.,CRC Practical Handbook of Biochemistry and Molecular Biology, CRC Press, Boca Raton, FL, pp. 3-70 [1989].
[cited by applicant]
Fowden, L., “Some observations on a hydroxypipecolic acid from thrift (Armeria maritima),” Biochem J., 70(4):629-33 [1958].
[cited by applicant]
Freier, S.M., et al., “Improved free-energy parameters for predictions of RNA duplex stability,” Proc. Natl. Acad. Sci USA, 83:9373-9377 (1986).
[cited by applicant]
Guo, Z., et al., “3′-End-Forming Signals of Yeast mRNA,” Mol. Cell. Biol., 15(11):5983-5990 [1995].
[cited by applicant]
Hara, R., “Characterization of novel 2-oxoglutarate dependent dioxygenases converting L-proline to cis-4-hydroxy+ proline,” Biochem Biophys Res Commun., 379(4):882-6 [2009].
[cited by applicant]
Henaut and Danchin in Neidhardt et al. [eds.], Escherichia coli and Salmonella, “Analysis and predictions from
[cited by applicant]
Henikoff, S., et al., “Amino acid substitution matrices from protein blocks,” Proc. Natl. Acad. Sci., 89:10915-10919 (1992).
[cited by applicant]
Jourdant, A., et al., “An efficient stereoselective and stereodivergent synthesis of (2R,3R)- and (2R,3S)-3-hydroxypipecolic acids,” Tetrahedron Lett., 41(36):7033-7036 [2000].
[cited by applicant]
Kalamkar, N.B., et al., “Chiron approach to the synthesis of (2S,3R)-3-hydroxypipecolic acid and (2R,3R)-3-hydroxy-2-hydroxymethylpiperidine from D-glucose,” J Org Chem., 73(9):3619-22 [2008].
[cited by applicant]
Kierzek, R., et al., “Polymer-Supported RNA Synthesis and Its Application to Test the Nearest-Neighbor Model for Duplex Stability,” Biochemistry, 25:7840-7846 (1986).
[cited by applicant]
Klein, C., et al., “A Simple Procedure for Selective Hydroxylation of I-Proline and I-Pipecolic Acid with Recombinantly Expressed ProlineHydroxylases,” Adv Synth. Catal., 353:1375-1383 [2011].
[cited by applicant]
Koszelewski, D., et al., “Immobilization of omega-transaminases by encapsulation in a sol-gel/celite matrix,” Journal of Molecular Catalysis B: Enzymatic, 63: 39-44 (2010).
[cited by applicant]
Kramer, B., et al., “Different base/base mismatches are corrected with different efficiencies by the methyl-directed DNA mismatch-repair system of
[cited by applicant]
Kumar, P., et al., “Asymmetric synthesis of both the enantiomers of trans-3-hydroxypipecolic acid,” J Org Chem., 70(1):360-3 [2005].
[cited by applicant]
Lawrence, C.C., et al., “Purification and initial characterization of proline 4-hydroxylase from Streptomyces griseoviridus P8648: a 2-oxoacid, ferrous-dependent dioxygenase involved in etamycin biosynthesis,” Biochem. …
[cited by applicant]
Lee, Y.K., et al., “The novel synthesis of two diastereomers of gamma-hydroxyproline,” Bull. Chem. Soc. Japan, 16:2924-26 [1973].
[cited by applicant]
Lemire, A., et al., “Stereoselective syntheses of L-pipecolic acid and (2S,3S)-3-hydroxypipecolic acid from a chiral N-imino-2-phenyl-1,2-dihydropyridine intermediate,” J Org Chem., 75(6):2077-80 [2010].
[cited by applicant]
Letavic, M.A., et al., “Synthesis and biological activity of selective pipecolic acid-based TNF-a converting enzyme (TACE) inhibitors,” Bioorg Medicinal Chem Lett., 12(10):1387-1390 [2002].
[cited by applicant]
Liang, N., et al., “Stereoselective total synthesis of cis- and trans-3-hydroxypipecolic acid,” J Org Chem., 70(24):10182-5 [2005].
[cited by applicant]
Ling, M., et al., “Approaches to DNA Mutagenesis:An Overview,” Anal. Biochem., 254:157-78 (1997).
[cited by applicant]
Majamaa, K., et al., “Differences between collagen hydroxylases and 2-oxoglutarate dehydrogenase in their inhibition by structural analogues of 2-oxoglutarate,” Biochem. J., 229:127-133 [1985].
[cited by applicant]
Marin, J., et al., “Synthesis of enantiopure 4-hydroxypipecolate and 4-hydroxylysine derivatives from a common 4,6-dioxopiperidinecarboxylate precursor,” J Org Chem., 69(1):130-41 [2004].
[cited by applicant]
Martin, A.R., et al., “Characterization of free and immobilized (S)-aminotransferase for acetophenone production,” Applied Microbiology and Biotechnology, 76(4): 843-851 (2007).
[cited by applicant]
Mateo, C., et al., “Epoxy sepabeads: a novel epoxy support for stabilization of industrial enzymes via very intense multipoint covalent attachment,” Biotechnology Progress 18(3):629-34 (2002).
[cited by applicant]
Matthes, H.W.D., et al., “Simultaneous rapid chemical synthesis of over one hundred oligonucleotides on a microscale,” EMBO J., 3(4):801-05 (1984).
[cited by applicant]
McInerney, J.O., “GCUA: general codon usage analysis,” Bioinformatics, 14(4):372-73, 1998.
[cited by applicant]
Minshull, J., et al., “Protein evolution by molecular breeding,” Curr. Op. Chem. Biol., 3(3):284-290 (1999).
[cited by applicant]
Mori, H., et al., “Detection of Novel Proline 3-Hydroxylase Activities in Streptomyces and Bacillus spp. by Regio- and Stereospecific Hydroxylation of I-Proline,” Appl. Environ. Microbiol., 62:1903-1907 [1996].
[cited by applicant]
Nakamura, Y., et al., “Codon usage tabulated from international DNA sequence databases: status for the year 2000,” Nucl. Acids Res., 28:292 [2000].
[cited by applicant]
Needleman, S., et al., “A general method applicable to the search for similarities in the amino acid sequence of two proteins,” J. Mol. Biol. 48:443-453 (1970).
[cited by applicant]
O'Connell, C.E., et al., “Synthesis and evaluation of some hydroxyproline-derived peptidomimetics as soprenyltransferase inhibitors” Chem Pharm Bull., 48(5):740-742 [2000].
[cited by applicant]
Pearson, W.R., “Improved tools for biological sequence comparison,” Proc. Nat'l. Acad. Sci. USA, 85:2444-2448 (1988).
[cited by applicant]
Petersen, L. et al., “Novel proline hydroxylase activities in the pneumocandin-producing fungus Glarea lozoyensis responsible for the formation of trans 3- and trans 4-hydroxyproline,” Appl Microbiol Biotechnol., 62(2-3…
[cited by applicant]
Ramaswarmy, S.G., et al., “One-vessel synthesis of 4-hydroxyproline from glyoxal and oxaloacetic acid,” J. Org. Chem., 42(21):3440-3443 [1977].
[cited by applicant]
Remuzon, P., “Trans-4-hydroxy-L-proline, a useful and versatile chiral starting block,” Tetrahedron, 52(44):13803-13835 [1996].
[cited by applicant]
Romanos, M.A., et al., “Foreign gene expression in yeast: a review,” Yeast 8:423-488 [1992].
[cited by applicant]
Romeo, J.T., et al., “CIS-4-Hydroxypipecolic Acid and 2,4-CIS-4,5-Trans-4,5-Dihydroxypipecolic Acid From Calliandra,” Phytochemistry 22(7):1615-1617 [1983].
[cited by applicant]
Rychlik, W., et al., “Optimization of the annealing temperature for DNA amplification in vitro,” Nucleic Acids Res, 18 (21):6409-6412 (1990).
[cited by applicant]
Shibasaki, T., et al., “Microbial Proline 4-Hydroxylase Screening and Gene Cloning,” Appl. Environ. Microbiol, 65(9):4028-31 [1999].
[cited by applicant]
Simonen, M., et al., “Protein Secretion in Bacillus Species,” Microbiological Reviews, 57:109-137 (1993).
[cited by applicant]
Smith, M., “In vitro mutagenesis,” Ann. Rev. Genet., 19:423-462 (1985).
[cited by applicant]
Smith, T., et al., “Comparison of Biosequences,” Adv. Appl. Math, 2:482-489 (1981).
[cited by applicant]
Stellwagen, E., “Dye Affinity Chromatography,” Current Protocols in Protein Science, Chapter 9, Unit 9.2-9.2.16 [2001].
[cited by applicant]
Stemmer, W., “DNA Shuffling by Random Fragmentation and Reassembly: In vitro Recombination for Molecular Evolution,” Proc. Natl. Acad. Sci. USA, 91:10747-10751 (1994).
[cited by applicant]
Stemmer, W.P.C., “Rapid evolution of a protein in vitro by DNA shuffling”, Nature, 370:389-391 (1994).
[cited by applicant]
Stenico, M., et al., “Codon usage in Caenorhabditis elegans: delineation of translational selection and mutational biases,” Nucl. Acids Res. 22(13):2437-46 [1994].
[cited by applicant]
Suggs, S.V., et al., “Use of synthetic oligodeoxyribonucleotides for the isolation of specific cloned DNA sequenes,” In Developmental Biology Using Purified Genes (Brown et al., eds.), pp. 683-693, Academic Press (1981).
[cited by applicant]
Tiwari, S., et al., “Prediction of probable genes by Fourier analysis of genomic sequences,” Comput. Appl. Biosci. 13(3):263-270 [1997].
[cited by applicant]
Truppo, M.D., et al., “Development of an Improved Immobilized CAL-B for the Enzymatic Resolution of a Key Intermediate to Odanacatib,” Organic Process Research & Development, 15:1033-1035 (2011).
[cited by applicant]
Uberbacher, E.C., et al., “Discovering and Understanding Genes in Human DNA Sequence Using GRAIL,” Methods Enzymol., 266:259-281 [1996].
[cited by applicant]
Vergnon, A.L., et al., “Solid phase synthesis of a library of hydroxyproline derivatives,” J Comb Chem., 6(1):91-8 [2004].
[cited by applicant]
Mlla-Komaroff, L., et al., “A bacterial clone synthesizing proinsulin,” Proc. Natl Acad. Sci. USA, 75:3727-3731 (1978).
[cited by applicant]
Wada, K., et al., “Codon usage tabulated from the GenBank genetic sequence data,” Nucl. Acids Res., 20:2111-2118 [1992].
[cited by applicant]
Wells, J.A., et al., “Cassette mutagenesis: an efficient method for generation of multiple mutations at defined sites,” Gene, 34:315-323 (1985).
[cited by applicant]
Wetmur, J. G., “DNA Probes: Applications of the Principles of Nucleic Acid Hybridization,” Crit Rev Biochem Mol Biol, 26(3/4):227-259 (1991).
[cited by applicant]
Wright, F., “The ‘effective number of codons’ used in a gene,” Gene 87:23-29 [1990].
[cited by applicant]
Yaegaki, K., et al., “Improved high-performance liquid chromatography method for quantitation of proline and hydroxyproline in biological materials,” J Chromatogr., 356(1):163-70 [1986].
[cited by applicant]
Yi, S., et al., “Covalent immobilization of omega-transaminase from Vibrio fluvialis JS17 on chitosan beads,” Process Biochemistry 42(5): 895-898 (2007).
[cited by applicant]
Zhang, J-H., et al., “Directed evolution of a fucosidase from a galactosidase by DNA shuffling and screening,”Proc. Nat. Acad. Sci., U.S.A., 94:4504-4509 (1997).
[cited by applicant]
Zhao, H., et al., “Molecular evolution by staggered extension process (StEP) in vitro recombination,” Nat. Biotechnol., 16:258-261 (1998).
[cited by applicant]
Chica, R.A., et al., “Semi-rational approaches to engineering enzyme activity: combining the benefits of directed evolution and rational design,” Current Opinion in Biotechnology, 16(4):378-384 [2005].
[cited by applicant]
Singh, R.K., et al., “Protein Engineering Approaches in the Post-Genomic Era,” Current Protein and Peptide Science, 18(4)1-11 [2017].
[cited by applicant]
Sen, S., et al., “Developments in Directed Evolution for Improving Enzyme Functions ,” Appl. Biochem. Biotech., 143(3):212-223 [2007].
[cited by applicant]
Accession No. Q92LF6 dated Dec. 1, 2001.
[cited by applicant]
Li, Y., et al., “Recent advances in engineering proteins for biocatalysis,” Biotechnology and Bioengineering, 111(7):1273-1287 [2014].
[cited by applicant]
Alignment to SEQ ID No. 226 of U.S. Pat. No. 9790527 dated Oct. 17, 2017.
[cited by applicant]
Koketsu, K., et al., “Refined Regio- and Stereoselective Hydroxylation of L-Pipecolic Acid by Protein Engineering of L-Proline cis-4-Hydroxylase Based on the X-ray Crystal Structure,” ACS Synthetic Biology, 4(4):383-392…
[cited by applicant]
Falcioni, F., et al., “Proline Availability Regulates Proline-4-Hydroxylase Synthesis and Substrate Update in Proline-Hydroxylating Recombinant
[cited by applicant]
16196205, Filing Receipt, Dec. 4, 2018.
[cited by applicant]
Genbank Accession No. CAC47686.1 dated Mar. 7, 2015.
[cited by applicant]