Skip to main content
Log in

Decrease of proteolytic degradation of recombinant hirudin produced by Pichia pastoris by controlling the specific growth rate

  • Published:
Biotechnology Letters Aims and scope Submit manuscript

Abstract

The effects of the specific growth rate and methanol concentration on the degradation of hirudin produced by recombinant Pichia pastoris were investigated. When a methanol-limited state and the specific growth rate of 0.02 h−1 were maintained during the fermentation, a minimum degradation of hirudin and a maximum specific hirudin production rate were achieved. By this strategy, the production of intact recombinant hirudin Hir65 reached 0.7 g l−1 in fed-batch fermentation. Its proportion was 35% to all forms of hirudin.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Similar content being viewed by others

References

  • Blank HW, Clark DS (1996) Biochemical Engineer. New York: Marcel Dekker, 702 pp.

    Google Scholar 

  • Cereghino JL, Cregg JM (2000) Heterologous protein expression in the methylotrophic yeast Pichia pastoris. FEMS Microbiol. Rev. 24: 45-66.

    Google Scholar 

  • Chang JY (1983) The functional domain of hirudin, a thrombinspecific inhibitor. FEBS Lett. 164: 307-313.

    Google Scholar 

  • d'Anjou MC, Daugulis AJ (2000) Mixed-feed exponential feeding for fed-batch culture of recombinant methylotrophic yeast. Biotechnol. Lett. 22: 341-346.

    Google Scholar 

  • d'Anjou MC, Daugulis AJ (2001) A rational approach to improving productivity in recombinant Pichia pastoris fermentation. Biotechnol. Bioeng. 72: 1-11.

    Google Scholar 

  • Heim J, Takabayashi K, Meyhack B, Marki W, Pohlig G (1994) C-terminal proteolytic degradation of recombinant desulfatohirudin and its mutants in the yeast Saccharomyces cerevisiae. Eur. J. Biochem. 226: 341-353.

    Google Scholar 

  • Katakura Y, Zhang WH, Zhang GQ, Omasa T, Kishimoto M, Goto YJ, 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: 482-87.

    Google Scholar 

  • Kim CH, Sohn JH, Choi ES, Rhee SK (1996) Effect of soybean oil on the enhanced expression of hirudin gene in Hansenula polymorpha. Biotechnol. Lett. 18: 417-422.

    Google Scholar 

  • Kobayashi K, Kuwae S, Ohya T, Ohda T, Ohyama M, Ohi H, Tomomitsu K, Ohmura T (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: 55-61.

    Google Scholar 

  • Rao KJ, Kim CH, Chung BH, Kim MK, Rhee SK (1999) Suppression of proteolytic degradation of recombinant hirudin from Saccharomyces cerevisiae using the O2-enriched air. Biotechnol. Lett. 21: 391-394.

    Google Scholar 

  • Sreekrishna K, Kropp KE (1996). Pichia pastoris. In: Wolf K, ed. Non-Conventional Yeast in Biotechnology: A Handbook. Berlin: Springer-Verlag, pp. 203-253.

    Google Scholar 

  • Sreekrishna K, Brankamp RG, Kropp KE, Blankenship DT, Tsay JT, Smith PL, Wierschke JD, Subramaniam A, Birkenberger LA (1997) Strategies for optimal synthesis and secretion of heterologous proteins in the methylotrophic yeast Pichia pastoris. Gene 190: 55-62.

    Google Scholar 

  • Zhang WH, Bevins MA, Plantz BA, Smith LA, Meager MM (2000) Modeling Pichia pastoris growth on methanol and optimization the production of a recombinant protein, the heavychain fragment C of botulinum neurotoxin, serotype A. Biotechnol. Bioeng. 70: 1-8.

    Google Scholar 

  • Zhou X-S, Lu J, Fan W-M, Zhang Y-X (2002) Development of a responsive methanol sensor and its application in Pichia pastoris fermentation. Biotechnol. Lett. 24: 643-646.

    Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Rights and permissions

Reprints and permissions

About this article

Cite this article

Zhou, XS., Zhang, YX. Decrease of proteolytic degradation of recombinant hirudin produced by Pichia pastoris by controlling the specific growth rate. Biotechnology Letters 24, 1449–1453 (2002). https://doi.org/10.1023/A:1019831406141

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1023/A:1019831406141

Navigation