Abstract
A recombinant strain of Pichia pastoris with a phenotype of MutS was used to produce angiostatin. Due to the low methanol consumption rate of this strain, both methanol and glycerol feedings, that produced oscillation in dissolved O2 concentration, were used during the expression phase to improve cell growth and angiostatin expression. However, enhanced cell growth led to nitrogen limitation that suppressed further production of angiostatin, but addition of ammonia allowed angiostatin concentration to reach 108 mg l−1 after an expression period of 96 h. The ratio of consumed glycerol to methanol of 1.5:1 (w/w) in the expression phase suggested that methanol played an important role in the metabolism of carbon sources.
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Brierley BA, Bussineau C, Kosson R, Melton A, Siegel RS (1990) Fermentation development of recombinant Pichia pastoris expressing the heterologous gene: bovine lysozyme. Ann. N.Y. Acad. Sci. 589: 350–362.
Chiruvolu V, Cregg JM, Meagher MM (1997) Recombinant protein production in an alcohol oxidase-defective strain of Pichia pastoris in fedbatch fermentations. Enzyme Microb. Technol. 21: 277–283.
Clare JJ, Romanos MA, Rayment FB, Rowedder JE, Smith MA, Payne MM, Sreekrishna K, Henwood CA (1991) Production of mouse epidermal growth factor in yeast: high-level secretion using Pichia pastoris strains containing multiple gene copies. Gene 105: 205–212.
Files D, Ogawa M, Scaman CH, Baldwin SA (2001) A Pichia pastoris fermentation process for producing high-levels of recombinant human cystatin-C. Enzyme Microb. Technol. 29: 335–340.
Katakura Y, Zhang W, Zhuang G, Omasa T, Kishmoto K, Goto Y, Suga KI (1998) Effect of methanol concentration on the production of human β2-glycoprotein I domain V by a recombinant Pichia pastoris: a simple system for the control of methanol concentration using a semiconductor gas sensor. J. Ferment. Bioeng. 86: 482–487.
Lin J, Panigraphy D, Trinh LB, Folkman J, Shiloach J (2000) Production process for recombinant human angiostatin in Pichia pastoris. J. Ind. Microbiol. Biotechnol. 24: 31–35.
Loewen MC, Liu X, Daugulis AJ (1997) Biosynthesis production of type II fish antifreeze protein: fermentation by Pichia pastoris. Appl. Microbiol. Biotechnol. 48: 480–486.
O'Reilly MS, Holmgren L, Shing Y, Chen C, Rosenthal RA, Moses M, Lane WS, Cao Y, Sage EH, Folkman J (1994) Angiostatin: a novel angiogenesis inhibitor that mediates the suppression of metastases by a Lewis lung carcinoma. Cell 79: 315–328.
Sim BKL, O'Reilly MS, Liang H, Fortier AH, He W, Madsen, JW, Lapcevich R, Nacy CA (1997) A recombinant human angiostatin protein inhibits experimental primary and matastatic cancer. Cancer Res. 57: 1329–1334.
Thorpe ED, d'Anjou MC, Daugulis AJ (1999) Sorbitol as a nonrepression carbon source for fed-batch fermentation of recombinant Pichia pastoris. Biotechnol. Lett. 21: 669–672.
Xin L, Zhang L, Xu R, Zhang Q, Ye Q, Li ZP, Gan RB (2001) Expression of human angiostatin in Pichia pastoris and the detection of its anti-angiogenic activity. Shengwu Huaxue Yu Shengwu Wuli Xuebao 33: 291–295 (in Chinese).
Zhang L, Ye Q, Xin L, Du P, Gan RB (2002) Effect of ammonium concentration on the growth of recombinant Pichia pastoris and expression of angiostatin. Weishengwuxue Tongbao 29: 23–26 (in Chinese).
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Xie, J., Zhang, L., Ye, Q. et al. Angiostatin production in cultivation of recombinant Pichia pastoris fed with mixed carbon sources. Biotechnology Letters 25, 173–177 (2003). https://doi.org/10.1023/A:1021905010021
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DOI: https://doi.org/10.1023/A:1021905010021