Chapter 11 : Heterologous Protein Expression in Yeasts and Filamentous Fungi

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Many filamentous fungi and yeast species have been used for heterologous protein. This chapter covers the most commonly used fungal expression systems (, , and species), with a focus on vector systems, promoter and leader sequences, posttranslational modifications, and fermentation scalability. Similar to other heterologous protein expression systems, proteolytic hydrolysis often causes instability of foreign proteins in . In comparison with and , filamentous fungi are less studied for heterologous protein production with respect to their transcription and translation control, mRNA stability, glycosylation, and regulation of protein secretion. There are several web-accessible organizations such as the American Type Culture Collection (ATCC), the (Fungal Genetics Stock Center (FGSC), the Agricultural Research Service Culture Collection (NRRL), and the Belgium Culture Collection (BCCP) from which the necessary tools for heterologous protein expression in filamentous fungi can be obtained. High mannose-type glycosylation is observed in filamentous fungi, but it is much less extensive than that seen in proteins produced in and species. Greater understanding of transcriptional control, mRNA stability, glycosylation, protein folding, and other posttranslational processes will allow us to overcome the difficulties associated with heterologous protein expression in fungi, such as hyperglycosylation in yeasts and poor secretion associated with some mammalian proteins. The exponential increase in fungal genomics and proteomics in recent years will no doubt fuel the advancement of filamentous fungi as heterologous protein expression hosts.

Citation: Zhang N, An Z. 2010. Heterologous Protein Expression in Yeasts and Filamentous Fungi, p 145-156. In Baltz R, Demain A, Davies J, Bull A, Junker B, Katz L, Lynd L, Masurekar P, Reeves C, Zhao H (ed), Manual of Industrial Microbiology and Biotechnology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816827.ch11
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1. Adams, B.,, D. J. Lowe,, A. T. Smith,, C. Scazzocchio,, S. Demais, and, R. C. Bray. 2002. Expression of Drosophila melanogaster xanthine dehydrogenase in Aspergillns nidulans and some properties of the recombinant enzyme. Biochem. J. 362: 223229.
2. Aleksenko, A., and , A. J. Clutterbuck. 1997. Autonomous plasmid replication in Aspergillus nidulans: AMA1 and MATE elements. Fungal Genet. Biol. 21: 373387.
3. Aleksenko, A., and, L. Ivanova. 1998. In vivo linearization and autonomous replication of plasmids containing human telomeric DNA in Aspergillus nidulans. Mol. Gen. Genet. 260: 159164.
4. Archer, D. B.,, D. J. Jeenes, and, D. A. Mackenzie. 1994. Strategies for improving heterologous protein production from filamentous fungi. Antonie van Leeuwenhoek 65: 245250.
5. Armstrong, K. A.,, T. Som,, F. C. Volkert,, A. Rose, and, J. R. Broach. 1989. Propagation and expression of genes in yeast using 2-micron circle vectors. Biotechnology 13: 165192.
6. Bae, J. Y.,, J. Laplaza, and, T. W. Jeffries. 2008. Effects of gene orientation and use of multiple promoters on the expression of XYL1 and XYL2 in Saccharomyces cerevisiae. Appl. Biochem. Biotechnol. 145: 6978.
7. Ballance, D. J.,, F. P. Buxton, and, G. Turner. 1983. Transformation of Aspergillus nidulans by the orotidine-5′-phosphate decarboxylase gene of Neurospora crassa. Biochem. Biophys. Res. Commun. 112: 284289.
8. Bautista, L. F.,, A. Aleksenko,, M. Hentzer,, A. Santerre-Henriksen, and, J. Nielsen. 2000. Antisense silencing of the creA gene in Aspergillus nidulans. Appl. Environ. Microbiol. 66: 45794581.
9. Berrin, J. G.,, W. R. McLauchlan,, P. Needs,, G. Williamson,, A. Puigserver,, P. A. Kroon, and, N. Juge. 2002. Functional expression of human liver cytosolic b-glucosidase in Pichia pastoris. Insights into its role in the metabolism of dietary glucosides. Eur. J. Biochem. 269: 249258.
10. Brake, A. J.,, J. P. Merryweather,, D. G. Coit,, U. A. Heberlein,, F. R. Masiarz,, G. T. Mullenbach,, M. S. Urdea,, P. Valenzuela, and, P. J. Barr. 1984. β-Factor-directed synthesis and secretion of mature foreign proteins in Saccharomyces cerevisiae. Proc. Natl. Acad. Sci. USA 81: 46424646.
11. Bretthauer, R. K., and, F. J. Castellino. 1999. Glycosylation of Pichia pastoris-derived proteins. Biotechnol. Appl. Biochem. 30: 193200.
12. Buckholz, R. G., and, M. A. Gleeson. 1991. Yeast systems for the commercial production of heterologous proteins. Biotechnology (New York) 9: 10671072.
13. Buxton, F. P.,, D. I. Gwynne, and, R. W. Davies. 1985. Transformation of Aspergillus niger using the argB gene of Aspergillus nidulans. Gene 37: 207214.
14. Cereghino, J. L., and, J. M. Cregg. 2000. Heterologous protein expression in the methylotrophic yeast Pichia pastoris. FEMS Microbiol. Rev. 24: 4566.
15. Chen, Y.,, D. Pioli, and, P. W. Piper. 1994. Overexpression of the gene for polyubiquitin in yeast confers increased secretion of a human leucocyte protease inhibitor. Biotechnology (New York) 12: 819823.
16. Chlebowicz-sledziewska, E., and, A. Z. Sledziewski. 1985. Construction of multicopy yeast plasmids with regulated centromere function. Gene 39: 2531.
17. Christensen, T.,, H. Woeldike,, E. Boel,, S. B. Mortensen,, K. Hjortshoej,, L. Thim, and, M. T. Hansen. 1988. High-level expression of recombinant genes in Aspergillus oryzae. Bio/Technology 6: 14191422.
18. Clarke, L., and, J. Carbon. 1980. Isolation of a yeast centromere and construction of functional small circular chromosomes. Nature 287: 504509.
19. Conesa, A.,, G. Weelink,, C. A. van den Hondel, and, P. J. Punt. 2001. C-terminal propeptide of the Caldariomyces fumago chloroperoxidase: an intramolecular chaperone? FEBS Lett. 503: 117120.
20. Cregg, J. M.,, J. L. Cereghino,, J. Shi, and, D. R. Higgins. 2000. Recombinant protein expression in Pichia pastoris. Mol. Biotechnol. 16: 2352.
21. Cregg, J. M.,, T. S. Vedvick, and, W. C. Raschke. 1993. Recent advances in the expression of foreign genes in Pichia pastoris. Biotechnology (New York) 11: 905910.
22. Day, D. A., and, M. F. Tuite. 1998. Post-transcriptional gene regulatory mechanisms in eukaryotes: an overview. J. Endocrinol. 157: 361371.
23. De nobel, J. G., and, J. A. Barnett. 1991. Passage of molecules through yeast cell walls: a brief essay-review. Yeast 7: 313323.
24. Devchand, M., and, D. I. Gwynne. 1991. Expression of heterologous proteins in Aspergillus. J. Biotechnol. 17: 39.
25. Felenbok, B.,, M. Flipphi, and, I. Nikolaev. 2001. Ethanol catabolism in Aspergillus nidulans: a model system for studying gene regulation. Prog. Nucleic Acid Res. Mol. Biol. 69: 149204.
26. Funk, M.,, R. Niedenthal,, D. Mumberg,, K. Brinkmann,, V. Ronicke, and, T. Henkel. 2002. Vector systems for heterologous expression of proteins in Saccharomyces cere-visiae. Methods Enzymol. 350: 248257.
27. Futcher, A. B., and, B. S. Cox. 1984. Copy number and the stability of 2-micron circle-based artificial plasmids of Saccharomyces cerevisiae. J. Bacteriol. 157: 283290.
28. Gardell, S. J.,, T. R. Hare,, J. H. Han,, H. Z. Markus,, B. J. Keech,, C. E. Carty,, R. W. Ellis, and, L. D. Schultz. 1990. Purification and characterization of human plasminogen activator inhibitor type I expressed in Saccharomyces cerevisiae. Arch. Biochem. Biophys. 278: 467474.
29. Gasser, B., and, D. Mattanovich. 2007. Antibody production with yeasts and filamentous fungi: on the road to large scale? Biotechnol. Lett. 29: 201212.
30. Gellissen, G. 2000. Heterologous protein production in methylotrophic yeasts. Appl. Microbiol. Biotechnol. 54: 741750.
31. Gietz, R. D., and, R. A. Woods. 2002. Transformation of yeast by lithium acetate/single-stranded carrier DNA/ polyethylene glycol method. Methods Enzymol. 350: 8796.
32. Giga-Hama, Y., and, H. Kumagai. 1999. Expression system for foreign genes using the fission yeast Schizosaccharomyces pombe. Biotechnol. Appl. Biochem. 30(Pt. 3) : 235244.
33. Goffeau, A.,, B. G. Barrell,, H. Bussey,, R. W. Davis,, B. Dujon,, H. Feldmann,, F. Galibert,, J. D. Hoheisel,, C. Jacq,, M. Johnston,, E. J. Louis,, H. W. Mewes,, Y. Murakami,, P. Philippsen,, H. Tettelin, and, S. G. Oliver. 2001. Life with 6000 genes. Science 274: 546, 563567.
34. Gouka, R. J.,, P. J. Punt,, J. G. Hessing, and, C. A. van den Hondel. 1996. Analysis of heterologous protein production in defined recombinant Aspergillus awamori strains. Appl. Environ. Microbiol. 62: 19511957.
35. Gouka, R. J.,, P. J. Punt, and, C. A. van den Hondel. 2001. Efficient production of secreted proteins by Aspergillus: progress, limitations and prospects. Appl. Microbiol. Biotechnol. 47: 111.
36. Gouka, R. J.,, P. J. Punt, and, C. A. van den Hondel. 1997. Glucoamylase gene fusions alleviate limitations for protein production in Aspergillus awamori at the transcriptional and (post) translational levels. Appl. Environ. Microbiol. 63: 488497.
37. Hackel, B. J.,, D. G. Huang,, J. C. Buboz,, X. X. Wang, and, E. V. Shusta. 2006. Production of soluble and active transferrin receptor-targeting single-chain antibody using Saccharomyces cerevisiae. Pharm. Res. 23: 790797.
38. Hamer, J. E., and, W. E. Timberlake. 1987. Functional organization of the Aspergillus nidulans trpC promoter. Mol. Cell. Biol. 7: 23522359.
39. Hamilton, S. R.,, R. C. Davidson,, N. Sethuraman,, J. H. Nett,, Y. W. Jiang,, S. Rios,, P. Bobrowicz,, T. A. Stadheim,, H. J. Li,, B. K. Choi,, D. Hopkins,, H. Wischnewski,, J. Roser,, T. Mitchell,, R. R. Strawbridge,, J. Hoopes,, S. Wildt, and, T. U. Gerngross. 2006. Humanization of yeast to produce complex terminally sialylated glycoproteins. Science 313: 14411443.
40. Hamilton, S. R., and, T. U. Gerngross. 2007. Glycosylation engineering in yeast: the advent of fully humanized yeast. Curr. Opin. Biotechnol. 18: 387392.
41. Harris, T. J., and, J. S. Emtage. 1986. Expression of heter-ologous genes in E. coli. Microbiol. Sci. 3: 2831.
42. Hasslacher, M.,, M. Schall,, M. Hayn,, R. Bona,, K. Rumbold,, J. Luckl,, H. Griengl,, S. D. Kohlwein, and, H. Schwab. 1997. High-level intracellular expression of hydroxynitrile lyase from the tropical rubber tree Hevea brasiliensis in microbial hosts. Protein Expr. Purif. 11: 6171.
43. Heinisch, J. J. 1993. Expression of heterologous phosphofructokinase genes in yeast. FEBS Lett. 328: 3540.
44. Hensing, M. C.,, R. J. Rouwenhorst,, J. J. Heijnen,, J. P. van Dijken, and, J. T. Pronk. 1995. Physiological and technological aspects of large-scale heterologous-protein production with yeasts. Antonie van Leeuwenhoek 67: 261279.
45. Hinnen, A.,, J. B. Hicks, and, G. R. Fink. 1978. Transformation of yeast. Proc. Natl. Acad. Sci. USA 75: 19291933.
46. Hitzeman, R. A.,, C. Y. Chen,, F. E. Hagie,, E. J. Patzer,, C. C. Liu,, D. A. Estell,, J. V. Miller,, A. Yaffe,, D. G. Kleid,, A. D. Levinson, and, H. Oppermann. 1983. Expression of hepatitis B virus surface antigen in yeast. Nucleic Acids Res. 11: 27452763.
47. Hitzeman, R. A.,, F. E. Hagie,, H. L. Levine,, D. V. Goeddel,, G. Ammerer, and, B. D. Hall. 1981. Expression of a human gene for interferon in yeast. Nature 293: 717722.
48. Hockney, R. C. 1994. Recent developments in heterologous protein production in Escherichia coli. Trends Biotechnol. 12: 456463.
49. Homann, M. J.,, W.-C. Suen,, N. Zhang, and, A. Zaks. 2006. Comparative analysis of chemical and biocatalytic syntheses of drug intermediates, p. 647–661. In R. Patel (ed.), Biocatalysis in the Pharmaceutical and Biotechnology Industries. Marcel Dekker, New York, NY.
50. Hong, F.,, N. Q. Meinander, and, L. J. Jonsson. 2002. Fermentation strategies for improved heterologous expression of laccase in Pichia pastoris. Biotechnol. Bioeng. 79: 438449.
51. Hottiger, T.,, J. Kuhla,, G. Pohlig,, P. Furst,, A. Spielmann,, M. Garn,, S. Haemmerli, and, J. Heim. 1995. 2-Micron vectors containing the Saccharomyces cerevisiae metallo-thionein gene as a selectable marker: excellent stability in complex media, and high-level expression of a recombinant protein from a CUP1-promoter-controlled expression cassette in cis. Yeast 11: 114.
52. Ishida, H.,, K. Matsumura,, Y. Hata,, A. Kawato,, K. Suginami,, Y. Abe,, S. Imayasu, and, E. Ishishima. 2001. Establishment of a hyper-protein production system in submerged Aspergillus oryzae culture under tyrosinase-encoding gene ( melO) promoter control. Appl. Microbiol. Biotechnol. 57: 131137.
53. Janowicz, Z. A.,, K. Melber,, A. Merckelbach,, E. Jacobs,, N. Harford,, M. Comberbach, and, C. P. Hollenberg. 1991. Simultaneous expression of the S and L surface antigens of hepatitis B, and formation of mixed particles in the methy-lotrophic yeast, Hansenula polymorpha. Yeast 7: 431443.
54. Jeenes, D. J.,, B. Marczinke,, D. A. MacKenzie, and, D. B. Archer. 1993. A truncated glucoamylase gene fusion for heterologous protein secretion from Aspergillus niger. FEMS Microbiol. Lett. 107: 267271.
55. Johnston, S. A.,, J. M. Salmeron, Jr., and, S. S. Dincher. 1987. Interaction of positive and negative regulatory proteins in the galactose regulon of yeast. Cell 50: 143146.
56. Karin, M.,, R. Najarian,, A. Haslinger,, P. Valenzuela,, J. Welch, and, S. Fogel. 1984. Primary structure and transcription of an amplified genetic locus: the CUP1 locus of yeast. Proc. Natl. Acad. Sci. USA 81: 337341.
57. Kariya, Y.,, K. Ishida,, Y. Tsubota,, Y. Nakashima,, T. Hirosaki,, T. Ogawa, and, K. Miyazaki. 2002. Efficient expression system of human recombinant laminin-5. J. Biochem. 132: 607612.
58. Kaslow, D. C., and, J. Shiloach. 1994. Production, purification and immunogenicity of a malaria transmission-blocking vaccine candidate: TBV25H expressed in yeast and purified using nickel-NTA agarose. Biotechnology (New York) 12: 494499.
59. Kelly, J. M., and, M. J. Hynes. 1985. Transformation of Aspergillus niger by the amdS gene of Aspergillus nidulans. EMBO J. 4: 475479.
60. Kingsman, S. M.,, A. J. Kingsman,, M. J. Dobson,, J. Mellor, and, N. A. Roberts. 1985. Heterologous gene expression in Saccharomyces cerevisiae. Biotechnol. Genet. Eng. Rev. 3: 377416.
61. Kitamoto, N.,, J. Matsui,, Y. Kawai,, A. Kato,, S. Yoshino,, K. Ohmiya, and, N. Tsukagoshi. 1998. Utilization of the TEF1-alpha gene ( TEF1) promoter for expression of polygalacturonase genes, pgaA and pgaB, in Aspergillus oryzae. Appl. Microbiol. Biotechnol. 50: 8592.
62. Kobayashi, K.,, S. Kuwae,, T. Ohya,, T. Ohda,, M. Ohyama,, H. Ohi,, K. Tomomitsu, and, T. Ohmura. 2000. High-level expression of recombinant human serum albumin from the methylotrophic yeast Pichia pastoris with minimal protease production and activation. J. Biosci. Bioeng. 89: 5561.
63. Kruszewska, J. S. 1999. Heterologous expression of genes in filamentous fungi. Acta Biochim. Pol. 46: 181195.
64. Kukuruzinska, M. A.,, M. L. Bergh, and, B. J. Jackson. 1987. Protein glycosylation in yeast. Annu. Rev. Biochem. 56: 915944.
65. Lai, M. T.,, D. Y. T. Liu, and, T. H. Hseu. 2007. Cell growth restoration and high level protein expression by the promoter of hexose transporter, HXT7, from Saccharomyces cerevisiae. Biotechnol. Lett. 29: 12871292.
66. Lau, W. W.,, K. R. Schneider, and , E. K. O’shea. 2001. A genetic study of signaling processes for repression of PHO5 transcription in Saccharomyces cerevisiae. Genetics 150: 13491359.
67. Lehle, L., and , E. Bause. 1984. Primary structural requirements for N-glycosylation and O-glycosylation of yeast mannoproteins. Biochim. Biophys. Acta 799: 246251.
68. Lemontt, J. F.,, C. M. Wei, and, W. R. Dackowski. 1985. Expression of active human uterine tissue plasminogen activator in yeast. DNA 4: 419428.
69. Lenouvel, F.,, P. J. van de Vondervoort, and, J. Visser. 2002. Disruption of the Aspergillus niger argB gene: a tool for transformation. Curr. Genet. 41: 425432.
70. Li, H.,, N. Sethuraman,, T. A. Stadheim,, D. Zha,, B. Prinz,, N. Ballew,, P. Bobrowicz,, B. K. Choi,, W. J. Cook,, M. Cukan,, N. R. Houston-cummings,, R. Davidson,, B. Gong,, S. R. Hamilton,, J. P. Hoopes,, Y. Jiang,, N. Kim,, R. Mansfield,, J. H. Nett,, S. Rios,, R. Strawbridge,, S. Wildt, and, T. U. Gerngross. 2006. Optimization of humanized IgGs in glycoengineered Pichia pastoris. Nat. Biotechnol. 24: 210215.
71. Li, M.,, F. Hubalek,, P. Newton-Vinson, and, D. E. Edmondson. 2002. High-level expression of human liver monoamine oxidase A in Pichia pastoris: comparison with the enzyme expressed in Saccharomyces cerevisiae. Protein Expr. Purif. 24: 152162.
72. Li, P.,, A. Anumanthan,, X. G. Gao,, K. Ilangovan,, V. V. Suzara,, N. Duzgunes, and, V. Renugopalakrishnan. 2007. Expression of recombinant proteins in Pichia pastoris. Appl. Biochem. Biotechnol. 142: 105124.
73. Lopes, T. S.,, J. Klootwijk,, A. E. Veenstra,, P. C. van der Aar,, H. van Heerikhuizen,, H. A. Raue, and , R. J. Planta. 1989. High-copy-number integration into the ribosomal DNA of Saccharomyces cerevisiae: a new vector for high-level expression. Gene 79: 199206.
74. Lubertozzi, D., and , J. D. Keasling. 2009. Developing Aspergillus as a host for heterologous expression. Biotechnol. Adv. 27: 5375.
75. Mach, H.,, D. B. Volkin,, R. D. Troutman,, B. Wang,, Z. Luo,, K. U. Jansen, and, L. Shi. 2006. Disassembly and reassembly of yeast-derived recombinant human papillomavirus viruslike particles (HPV VLPs). J. Pharm. Sci. 95: 21952206.
76. Madzak, C.,, M. C. Mimmi,, E. Caminade,, A. Brault,, S. Baumberger,, P. Briozzo,, C. Mougin, and , C. Jolivalt. 2006. Shifting the optimal pH of activity for a laccase from the fungus Trametes versicolor by structure-based mutagenesis. Protein Eng. Des. Sel. 19: 7784.
77. Madzak, C.,, L. Otterbein,, M. Chamkha,, S. Moukha,, M. Asther,, C. Gaillardin, and, J. M. Beckerich. 2005. Heterologous production of a laccase from the basidiomycete Pycnoporus cinnabarinus in the dimorphic yeast Yarrowia lipolytica. FEMS Yeast Res. 5: 635646.
78. Maras, M.,, I. van Die,, R. Contreras, and , C. A. van den Hondel. 2001. Filamentous fungi as production organisms for glycoproteins of bio-medical interest. Glycoconj. J. 16: 99107.
79. Martinez-Ruiz, A.,, A. Martinez del Pozo,, J. Lacadena,, J. M. Mancheno,, M. Onaderra,, C. Lopez-Otin, and, J. G. Gavilanes. 1998. Secretion of recombinant pro- and mature fungal alpha-sarcin ribotoxin by the methylotrophic yeast Pichia pastoris: the Lys-Arg motif is required for maturation. Protein Expr. Purif. 12: 315322.
80. Matsuyama, A.,, A. Shirai, and, M. Yoshida. 2008. A series of promoters for constitutive expression of heterologous genes in fission yeast. Yeast 25: 371376.
81. Mattern, I. E.,, S. Unkles,, J. R. Kinghorn,, P. H. Pouwels, and , C. A. van den Hondel. 1987. Transformation of Aspergillus oryzae using the A. niger pyrG gene. Mol. Gen. Genet. 210: 460461.
82. Mccarthy, J. E. 1998. Posttranscriptional control of gene expression in yeast. Microbiol. Mol. Biol. Rev. 62: 14921553.
83. Mendoza-Vega, O.,, J. Sabatie, and, S. W. Brown. 1994. Industrial production of heterologous proteins by fed-batch cultures of the yeast Saccharomyces cerevisiae. FEMS Microbiol. Rev. 15: 369410.
84. Moir, D. T., and , D. R. Dumais. 1987. Glycosylation and secretion of human alpha-1-antitrypsin by yeast. Gene 56: 209217.
85. Morawski, B.,, Z. Lin,, P. Cirino,, H. Joo,, G. Bandara, and, F. H. Arnold. 2000. Functional expression of horseradish peroxidase in Saccharomyces cerevisiae and Pichia pastoris. Protein Eng. 13: 377384.
86. Murray, A. W., and , J. W. Szostak. 1983. Pedigree analysis of plasmid segregation in yeast. Cell 34: 961970.
87. Nevalainen, K. M.,, V. S. Te’o, and, P. L. Bergquist. 2005. Heterologous protein expression in filamentous fungi. Trends Biotechnol. 23: 468474.
88. Nikolaev, I.,, M. Mathieu,, P. van de Vondervoort,, J. Visser, and, B. Felenbok. 2002. Heterologous expression of the Aspergillus nidulans alcR-alcA system in Aspergillus niger. Fungal Genet. Biol. 37: 8997.
89. Nyyssonen, E.,, M. Penttila,, A. Harkki,, A. Saloheimo,, J. K. Knowles, and, S. Keranen. 1993. Efficient production of antibody fragments by the filamentous fungus Trichoderma reesei. Biotechnology (New York) 11: 591595.
90. O’callaghan, J.,, M. M. O’Brien,, K. Mcclean, and, A. D. Dobson. 2002. Optimisation of the expression of a Trametes versicolor laccase gene in Pichia pastoris. J. Ind. Microbiol. Biotechnol. 29: 5559.
91. Outchkourov, N. S.,, W. J. Stiekema, and, M. A. Jongsma. 2002. Optimization of the expression of equistatin in Pichia pastoris. Protein Expr. Purif. 24: 1824.
92. Papagianni, M.,, N. Joshi, and , M. Moo-Young. 2001. Comparative studies on extracellular protease secretion and glucoamylase production by free and immobilized Aspergillus niger cultures. J. Ind. Microbiol. Biotechnol. 29: 259263.
93. Park, M. A.,, H. J. Kim, and, H. J. Kim. 2008. Optimum conditions for production and purification of human papillomavirus type 16 L1 protein from Saccharomyces cerevisiae. Protein Expr. Purif. 59: 175181.
94. Paus, E. J.,, J. Willey,, R. J. Ridge,, C. R. Legg,, M. A. Finkelman,, T. J. Novitsky, and, P. A. Ketchum. 2002. Production of recombinant endotoxin neutralizing protein in Pichia pastoris and methods for its purification. Protein Expr. Purif. 26: 202210.
95. Pohlig, G.,, W. Zimmermann, and, J. Heim. 1991. Influence of yeast proteases on hirudin expression in Saccharo-myces cerevisiae. Biomed. Biochim. Acta 50: 711716.
96. Porres, J. M.,, M. J. Benito, and, X. G. Lei. 2002. Functional expression of keratinase ( kerA) gene from Bacillus licheniformis in Pichia pastoris. Biotechnol. Lett. 24: 631636.
97. Price, V. L.,, W. E. Taylor,, W. Clevenger,, M. Worthing-ton, and, E. T. Young. 1990. Expression of heterologous proteins in Saccharomyces cerevisiae using the ADH2 promoter. Methods Enzymol. 185: 308318.
98. Proudfoot, A. E.,, G. Turcatti,, T. N. Wells,, M. A. Payton, and, D. J. Smith. 1994. Purification, cDNA cloning and heterologous expression of human phosphomannose isomerase. Eur. J. Biochem. 219: 415423.
99. Punt, P. J.,, M. A. Dingemanse,, A. Kuyvenhoven,, R. D. M. Soede,, P. H. Pouwels, and , C. A. van den Hondel. 1992. Functional elements in the promoter region of the Aspergillus nidulans gpdA gene encoding glyceraldehyde-3-phosphate dehydrogenase. Gene 93: 101109.
100. Punt, P. J.,, R. P. Oliver,, M. A. Dingemanse,, P. H. Pouwels, and , C. A. van den Hondel. 2001. Transformation of Aspergillus based on the hygromycin B resistance marker from Escherichia coli. Gene 56: 117124.
101. Punt, P. J.,, N. van Biezen,, A. Conesa,, A. Albers,, J. Mangnus, and , C. van den Hondel. 2001. Filamentous fungi as cell factories for heterologous protein production. Trends Biotechnol. 20: 200206.
102. Punt, P. J., and , C. A. van den Hondel. 2001. Transformation of filamentous fungi based on hygromycin B and phleo-mycin resistance markers. Methods Enzymol. 216: 447457.
103. Punt, P. J.,, N. D. Zegers,, M. Busscher,, P. H. Pouwels, and , C. A. van den Hondel. 2001. Intracellular and extracellular production of proteins in Aspergillus under the control of expression signals of the highly expressed Aspergillus nidulans gpdA gene. J. Biotechnol. 17: 1933.
104. Ramqvist, T.,, K. Andreasson, and, T. Dalianis. 2007. Vaccination, immune and gene therapy based on viruslike particles against viral infections and cancer. Expert Opin. Biol. Ther. 7: 9971007.
105. Raymond, C. K.,, T. Bukowski,, S. D. Holderman,, A. F. Ching,, E. Vanaja, and, M. R. Stamm. 1998. Development of the methylotrophic yeast Pichia methanolica for the expression of the 65 kilodalton isoform of human glutamate decarboxylase. Yeast 14: 1123.
106. Record, E.,, P. J. Punt,, M. Chamkha,, M. Labat,, C. A. van den Hondel, and, M. Asther. 2002. Expression of the Pycnoporus cinnabarinus laccase gene in Aspergillus niger and characterization of the recombinant enzyme. Eur. J. Biochem. 269: 602609.
107. Reddy, S. T., and , N. M. Dahms. 2002. High-level expression and characterization of a secreted recombinant cation-dependent mannose 6-phosphate receptor in Pichia pastoris. Protein Expr. Purif. 26: 290300.
108. Romanos, M. A.,, C. A. Scorer, and, J. J. Clare. 1992. Foreign gene expression in yeast: a review. Yeast 8: 423488.
109. Rothstein, R. J. 1983. One-step gene disruption in yeast. Methods Enzymol. 101: 202211.
110. Russell, C.,, J. Mawson, and, P. L. Yu. 1991. Production of recombinant products in yeasts: a review. Aust. J. Biotechnol. 5: 4855.
111. Sakai, A.,, F. Ozawa,, T. Higashizaki,, Y. Shimizu, and, F. Hishinuma. 1991. Enhanced secretion of human nerve growth factor from Saccharomyces cerevisiae using an advanced delta-integration system. Biotechnology (New York) 9: 13821385.
112. Sakai, Y.,, T. Rogi,, R. Takeuchi,, N. Kato, and, Y. Tani. 1995. Expression of Saccharomyces adenylate kinase gene in Candida boidinii under the regulation of its alcohol oxidase promoter. Appl. Microbiol. Biotechnol. 42: 860864.
113. Sarramegna, V.,, F. Talmont,, M. Seree de Roch,, A. Milon, and, P. Demange. 2002. Green fluorescent protein as a reporter of human μ-opioid receptor overexpression and localization in the methylotrophic yeast Pichia pastoris. J. Biotechnol. 99: 2339.
114. Schilling, B. M.,, J. C. Goodrick, and, N. C. Wan. 2001. Scale-up of a high cell density continuous culture with Pichia pastoris X-33 for the constitutive expression of rhchitinase. Biotechnol. Prog. 17: 629633.
115. Schultz, L. D.,, H. Z. Markus,, K. J. Hofmann,, D. L. Montgomery,, C. T. Dunwiddie,, P. J. Kniskern,, R. B. Freedman,, R. W. Ellis, and, M. F. Tuite. 1994. Using molecular genetics to improve the production of recombinant proteins by the yeast Saccharomyces cerevisiae. Ann. N. Y. Acad. Sci. 721: 148157.
116. Schuster, E.,, N. Dunn-coleman,, J. C. Frisvad, and , P. W. M. van Dijck. 2001. On the safety of Aspergillus niger: a review. Appl. Microbiol. Biotechnol. 59: 426435.
117. Shen, S.,, G. Sulter,, T. W. Jeffries, and, J. M. Cregg. 1998. A strong nitrogen source-regulated promoter for controlled expression of foreign genes in the yeast Pichia pastoris. Gene 216: 93102.
118. Shiba, Y.,, C. Ono,, F. Fukui,, I. Watanabe,, N. Serizawa,, K. Gomi, and, H. Yoshikawa. 2001. High-level secretory production of phospholipase A1 by Saccharomyces cerevisiae and Aspergillus oryzae. Biosci. Biotechnol. Biochem. 65: 94101.
119. Shuster, J. R.,, D. Moyer, and, H. Lee. 1992. A DNA vector system for the integration and amplification of heterologous DNA into dispersed chromosomal locations in the yeast, Saccharomyces cerevisiae, genome. Abstr. Pap. Am. Chem. Soc. 203:1198.
120. Sleep, D.,, G. P. Belfield, and, A. R. Goodey. 1990. The secretion of human serum albumin from the yeast Saccharomyces cerevisiae using five different leader sequences. Biotechnology (New York) 8: 4246.
121. Smith, J. D., and , A. S. Robinson. 2002. Overexpression of an archaeal protein in yeast: secretion bottleneck at the ER. Biotechnol. Bioeng. 79: 713723.
122. Stepien, P. P.,, R. Brousseau,, R. Wu,, S. Narang, and, D. Y. Thomas. 1983. Synthesis of a human insulin gene. VI. Expression of the synthetic proinsulin gene in yeast. Gene 24: 289297.
123. Tucker, M., and , R. Parker. 2000. Mechanisms and control of mRNA decapping in Saccharomyces cerevisiae. Annu. Rev. Biochem. 69: 571595.
124. Tuite, M. F.,, M. J. Dobson,, N. A. Roberts,, R. M. King,, D. C. Burke,, S. M. Kingsman, and , A. J. Kingsman. 1982. Regulated high efficiency expression of human interferon-alpha in Saccharomyces cerevisiae. EMBO J. 1: 603608.
125. Unkles, S. E.,, E. I. Campbell,, P. J. Punt,, K. L. Hawker,, R. Contreras,, A. R. Hawkins,, C. A. Van den Hondel, and, J. R. Kinghorn. 1992. The Aspergillus niger niaD gene encoding nitrate reductase: upstream nucleotide and amino acid sequence comparisons. Gene 111: 149155.
126. Valenzuela, P.,, A. Medina, and, W. J. Rutter. 1982. Synthesis and assembly of hepatitis-B virus surface-antigen particles in yeast. Nature 298: 347350.
127. van den Hombergh, J. P.,, P. J. van de Vondervoort,, L. Fraissinet-Tachet, and, J. Visser. 1997. Aspergillus as a host for heterologous protein production: the problem of proteases. Trends Biotechnol. 15: 256263.
128. Vasavada, A. 1995. Improving productivity of heterologous proteins in recombinant Saccharomyces cerevisiae fermentations. Adv. Appl. Microbiol. 41: 2554.
129. Wang, L.,, D. Ridgway,, T. Gu, and , M. Moo-Young. 2005. Bioprocessing strategies to improve heterologous protein production in filamentous fungal fermentations. Biotechnol. Adv. 23: 115129.
130. Wang, S. H.,, T. S. Yang,, S. M. Lin,, M. S. Tsai,, S. C. Wu, and, S. J. Mao. 2002. Expression, characterization, and purification of recombinant porcine lactoferrin in Pichia pastoris. Protein Expr. Purif. 25: 4149.
131. Ward, M.,, C. Lin,, D. C. Victoria,, B. P. Fox,, J. A. Fox,, D. L. Wong,, H. J. Meerman,, J. P. Pucci,, R. B. Fong,, M. H. Heng,, N. Tsurushita,, C. Gieswein,, M. Park, and, H. Wang. 2004. Characterization of humanized antibodies secreted by Aspergillus niger. Appl. Environ. Microbiol. 70: 25672576.
132. Ward, P. P.,, J. Y. Lo,, M. Duke,, G. S. May,, D. R. Headon, and, O. M. Conneely. 1992. Production of biologically active recombinant human lactoferrin in Aspergillus oryzae. Biotechnology (New York) 10: 784789.
133. Ward, P. P.,, G. S. May,, D. R. Headon, and, O. M. Conneely. 1992. An inducible expression system for the production of human lactoferrin in Aspergillus nidulans. Gene 122: 219223.
134. Wilcox, G., and , G. M. Studnicka. 1988. Expression of foreign proteins in microorganisms. Biotechnol. Appl. Biochem. 10: 500509.
135. Woo, M. K.,, J. M. An,, J. D. Kim,, S. N. Park, and, H. J. Kim. 2008. Expression and purification of human papillomavirus 18 L1 virus-like particle from Saccharomyces cerevisiae. Arch. Pharmacal Res. 31: 205209.
136. Xu, P.,, D. Raden,, F. J. Doyle, and, A. S. Robinson. 2005. Analysis of unfolded protein response during single-chain antibody expression in Saccharomyces cerevisiae reveals different roles for BiP and PDI in folding. Metab. Eng. 7: 269279.
137. Yelton, M. M.,, J. E. Hamer, and, W. E. Timberlake. 1984. Transformation of Aspergillus nidulans by using a trpC plasmid. Proc. Natl. Acad. Sci. USA 81: 14701474.
138. Zhang, N.,, D. L. Daubaras, and , W.-c. Suen. 2001. Heterologous protein expression in yeasts and filamentous fungi, p. 667–687. In Z. An (ed.), Handbook of Industrial Mycology. Marcel Dekker, New York, NY.


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Pros and cons for some commonly used heterologous protein expression systems

Citation: Zhang N, An Z. 2010. Heterologous Protein Expression in Yeasts and Filamentous Fungi, p 145-156. In Baltz R, Demain A, Davies J, Bull A, Junker B, Katz L, Lynd L, Masurekar P, Reeves C, Zhao H (ed), Manual of Industrial Microbiology and Biotechnology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816827.ch11
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Vector systems used for heterologous protein expression in

Citation: Zhang N, An Z. 2010. Heterologous Protein Expression in Yeasts and Filamentous Fungi, p 145-156. In Baltz R, Demain A, Davies J, Bull A, Junker B, Katz L, Lynd L, Masurekar P, Reeves C, Zhao H (ed), Manual of Industrial Microbiology and Biotechnology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816827.ch11
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Selectable markers used for heterologous gene expression in species

Citation: Zhang N, An Z. 2010. Heterologous Protein Expression in Yeasts and Filamentous Fungi, p 145-156. In Baltz R, Demain A, Davies J, Bull A, Junker B, Katz L, Lynd L, Masurekar P, Reeves C, Zhao H (ed), Manual of Industrial Microbiology and Biotechnology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816827.ch11
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Promoters used in heterologous gene expression in species

Citation: Zhang N, An Z. 2010. Heterologous Protein Expression in Yeasts and Filamentous Fungi, p 145-156. In Baltz R, Demain A, Davies J, Bull A, Junker B, Katz L, Lynd L, Masurekar P, Reeves C, Zhao H (ed), Manual of Industrial Microbiology and Biotechnology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816827.ch11

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