title: Overproduction of 15N-labeled r-RGD-hirudin in pichia pastoris for NMR studies.
abstract: The novel recombinant hirudin, r-RGD-hirudin, inhibits thrombin and platelet aggregation. Here, we reported over-expression of (15)N-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 culture in a fermenter. The purified (15)N-labeled r-RGD-hirudin retained full biological activity and was uniformly labeled. Heteronuclear NMR of the (15)N-labeled r-RGD-hirudin was performed for the first time, and all signals in the heteronuclear single quantum coherence (HSQC) spectrum were successfully assigned.
authors.affiliation: Center of Analysis and Measurement, Fudan University, Shanghai 200433, China.
authors.forename: Jue
authors.initials: J
authors.lastname: Wang
authors.forename: Yanling
authors.initials: Y
authors.lastname: Zhang
authors.forename: Shiyun
authors.initials: S
authors.lastname: Li
authors.forename: Xingang
authors.initials: X
authors.lastname: Liu
authors.forename: Xiaomin
authors.initials: X
authors.lastname: Yan
authors.forename: Houyan
authors.initials: H
authors.lastname: Song
authors.forename: Min
authors.initials: M
authors.lastname: Yu
authors.forename: Linsen
authors.initials: L
authors.lastname: Dai
authors.forename: Wei
authors.initials: W
authors.lastname: Mo
mesh_terms: D001709:Biotechnology / Q000379:methods*; D004591:Electrophoresis, Polyacrylamide Gel; D006629:Hirudins / Q000235:genetics / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D009587:Nitrogen Isotopes; D010843:Pichia / Q000378:metabolism; D015854:Up-Regulation
chemical_list: D006629:Hirudins; D009587:Nitrogen Isotopes; C544959:RGD-hirudin