Pichia pastoris

Until 1987, the (R)-PaHNL from almonds was the only HNL used as catalyst in the enantioselective preparation of cyanohydrins. Therefore, it was of great interest to get access to HNLs which catalyze the formation of (5 )-cyanohydrins. (5 )-SbHNL [EC 4.1.2.11], isolated from Sorghum bicolor, was the first HNL used for the preparation of (5 )-cyanohydrins. Since the substrate range of SbHNL is limited to aromatic and heteroaromatic aldehydes as substrates, other enzymes with (5 )-cyanoglycosides have been investigated as catalysts for the synthesis of (5 )-cyanohydrins. The (5 )-HNLs from cassava (Manihot esculenta, MeHNL) and from Hevea brasiliensis (HbHNL) proved to be highly promising candidates for the preparation of (5 )-cyanohydrins. Both MeHNL and HbHNL have been overexpressed successfully in Escherichia coli, Saccharomyces cerevisiae and Pichia pastoris. 

R)- and ( -selective HNLs. A number of recombinant HNLs have also been expressed in E. coli, Saccharomyces cerevisiae, and Pichia pastoris. Recently, protein engineering has been successfully applied to the development of a tailor-made HNL for large-scale production of specific cyanohydrins [69,70]. 

Macauley-Patrick, S., Fazenda, M.L., McNeil, B. and Harvey, L.M. (2005) Heterologous protein production using the Pichia pastoris expression system. Yeast, 22, 249-270. 

Chen, Y.-R., Huang, H.-H., Cheng, Y.-F. et al. (2006) Expression of a cholesterol oxidase gene from Arthrobacter simplex in Escherichia coli and Pichia pastoris. Enzyme and Microbial Technology, 39, 854-860. 

Traditional and well-established yeast species are Saccharomyces cerevisiae, Hansenula polymorpha, Klyveromyces lactis, Pichia pastoris and Schizosaccharomyces pombe. With every year that passes they are increasingly being used in industrial and pharmaceutical enzyme production on a large scale. Many further yeasts present interesting features (e.g. Arxula adeninivorans and Yarrowia lipolytica), but are not that widely used. 

Guoa, M., Hang, H. and Zhua, T. (2008) Effect of glycosylation on biochemical characterization of recombinant phytase expressed in Pichia pastoris. Enzyme and Microbial Technology, 42, 340-345. 

Sletta, H., Aune, R., Nedal, A. etal. (2007) The presence of N-terminal secretion signal sequences leads to strong stimulation of the total expression levels of three tested medically important proteins during high-cell-density cultivations of Pichia pastoris. Applied and Environmental Microbiology, 73 (3), 906-912. 

Wu, S., Fallon, R.D. and Payne, M.S. (1997) Over-production of stereoselective nitrile hydratase from Pseudomonas putida 5B in Pichia pastoris activity requires a novel downstream protein. Applied Microbiology and Biotechnology, 48 (6), 704—708. 

Choi, D.H. and Keum, K.C. (2006) Production of recombinant proteins by high cell density culture of Pichia pastoris. Chemical Engineering Science, 61, 876-885. 

Baneyx, F. I 491 Rcconibi nail protein expression in Pichia pastoris. Current Opinion in Biotechnology, 10(5), 411—421. 

Sorensen, H.P. and Mortensen, K.K. (2005) Advanced genetic strategies for recombinant protein expression in Pichia pastoris. Journal of Biotechnology, 115 (2), 113-128. 

Swartz, J.R. (2001) Advances in Pichia pastoris production of therapeutic proteins. Current Opinion in Biotechnology, 12 (2), 195-201. 

Choi, J.H. and Lee, S.Y. (2004) Secretory and extracellular production of recombinant proteins using Pichia pastoris. Applied Microbiology and Biotechnology, 64 (5), 625-635. 

Misawa, S. and Kumagai, I. (1999) Refolding of therapeutic proteins produced in Pichia pastoris as inclusion bodies. Biopolymers, 51 (4), 297-307. 

Shokri, A., Sanden, A.M. and Larsson, G. (2003) Cell and process design for targeting of recombinant protein into the culture medium of Pichia pastoris. Applied Microbiology and Biotechnology, 60 (6), 654-664. 

Trimble, R.B., Atkinson, P.H., Tschopp, J.F. et al. (1991) Structure of oligosaccharides on Saccharomyces SUC2 invertase secreted by the methylotrophic yeast Pichia pastoris. The Journal of Biological Chemistry, 266 (34), 22807-22817. 

Weis, R., Luiten, R., Skranc, W. et al. (2004) Reliable high-throughput screening with Pichia pastoris by limiting yeast cell death phenomena. FEMS Yeast Research, 5 (2), 179-189. 

Cereghino, J.L. and Cregg, J.M. (2000) Heterologous protein expression in the methylotrophic yeast Pichia pastoris. FEMS Microbiology Reviews, 24 (1), 45-66. 

Gellissen, G., Kunze, G., Gaillardin, C. et al. (2005) New yeast expression platforms based on methylotrophic Hansenula polymorpha and Pichia pastoris and on dimorphic Arxula adeninivorans and Yarrowia lipolytica - a comparison. FEMS Yeast Research, 5 (11), 1079-1096. 

Lin-Cereghino, J., Wong, W.W., Xiong, S. et al. (2005) Condensed protocol for competent cell preparation and transformation of the methylotrophic yeast Pichia pastoris. Biotechniques, 38 (1), 44, 46, 48. 

Engelking, H., Pfaller, R., Wich, G. and Weuster-Botz, D. (2004) Stereoselective reduction of ethyl 4-chloro acetoacetate with recombinant Pichia pastoris. Tetrahedron Asymmetry, 15 (22), 3591-3593. 

Dienys, G., Jarmalavicius, S., Budriene, S. et al. (2003) Alcohol oxidase from the yeast Pichia pastoris — a potential catalyst for organic synthesis. Journal of Molecular Catalysis B-Enzymatic, 21 (1-2), 47—49. 

Hermann, M., Kietzmann, M.U., Ivancic, M. et al. (2008) Alternative pig liver esterase (APLE) - cloning, identification and functional expression in Pichia pastoris of a versatile new biocatalyst. Journal of Biotechnology, 133 (3), 301-310. 

Vuorela, A., Myllyharju, J., Nissi, R., Pihlajaniemi, T. and Kivirikko, K.I. (1997) Assembly of human prolyl 4-hydroxylase and type III collagen in the yeast Pichia pastoris. formation of a stable enzyme tetramer requires coexpression... 

Doring, F., S. Theis, and H. Daniel. Expression and functional characterization of the mammalian intestinal peptide transporter PepTl in the methylotropic yeast Pichia pastoris. Biochem. Biophys. Res. Commun. 1997, 232, 656-662. 

Kawakami, K. and Furukawa, S.Y. (1997) Alcohol-oxidation activity of whole cells of Pichia pastoris entrapped in hybrid gels composed of Ca-alginate and organic silicate. Applied Microbiology and Biotechnology, 67, 23-31. 

Allerson, C.R., Martinez, A., Yikilmaz, E., and Rouault, T.A. (2003) A high-capacity RNA affinity column for the purification of human IRP1 and IRP2 overexpressed in Pichia pastoris. RNA 9, 364. 

Although proteins can be expressed in many heterologous production systems, including bacteria such as Proteus mirabilis [1], fungi such as Pichia pastoris [2, 3] and Aspergillus awamori [4] and insect cells [5, 6], the pharmaceutical industry has narrowed down process development to a small number of platform technologies ... 

ANDERSEN, M.D., M0LLER, B.L., Cytochromes P450 from Cassava (Manihot esculenta Crantz) catalyzing the first steps in the biosynthesis of the cyanogenic glucosides linamarin and lotaustralin cloning, functional expression in Pichia pastoris and substrate specificity of the isolated recombinant enzymes, J. Biol. Chem., 2000,275, 1966-1975. 

Kristensen AK, Brunstedt J, Nielsen JE, Mikkelsen JD, Roepsstorff P, Nielsen KK. Processing, disulfide pattern and biological activity of sugar beet defensin AX2, expression in Pichia pastoris. Protein Expr Purif 1999 16 377-387. 

Pakkanen, O., Hamalainen, E. R., Kivirikko, K. I., and Myllyharju.J. (2003). Assembly of stable human type I and III collagen molecules from hydroxylated recombinant chains in the yeast Pichia pastoris. Effect of an engineered C-terminal oligomerization domain foldon./. Biol. Chem. 278, 32478-32483. 

Fig. 18.—C-l Regions of 13C-N.m.r. Spectra of a,)3-D-Mannopyranans from Pichia pastoris (A) and Citeromyces matritensis (B), and Those of Oligosaccharides Formed by Partial Acetolysis (C and D, Respectively). (Solvent, DjO temperature, 70° chemical shifts expressed as 8C, relative to external tetramethylsilane.)...

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