Overproduction of 15N-Labeled r-RGD-Hirudin in Pichia pastoris for NMR Studies

Author(s): Jue Wang, Yanling Zhang, Shiyun Li, Xingang Liu, Xiaomin Yan, Houyan Song, Min Yu, Linsen Dai, Wei Mo

Journal Name: Protein & Peptide Letters

Volume 17 , Issue 10 , 2010

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The novel recombinant hirudin, r-RGD-hirudin, inhibits thrombin and platelet aggregation. Here, we reported over-expression of 15N-labeled r-RGD-hirudin by Pichia pastoris in minimal medium. After extensive optimization, the yield of active r-RGD-hirudin reached ~600 mg/L when the yeast cells were cultured in a fermenter. The purified 15N-labeled r-RGD-hirudin retained full biological activity and was uniformly labeled. Heteronuclear NMR of the 15N-labeled r-RGD-hirudin was performed for the first time, and all signals in the heteronuclear single quantum coherence (HSQC) spectrum were successfully assigned.

Keywords: 15N-Labeling, NMR, Pichia pastoris, r-RGD-Hirudin

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Article Details

Year: 2010
Page: [1228 - 1233]
Pages: 6
DOI: 10.2174/092986610792231465
Price: $65

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