|
|
Construction and Expression of sGLP-1- HSA Fusion protein |
GAO Hua1 XU Chong-bo1 LI Yuan2 |
1.Key Laboratory of Bioorganic Chemistry of Liaoning Province|College of Medicine| Dalian University|Dalian116622|China 2.Dalian D.N Bioengineering CO.|LTD|Dalian116600|China |
|
|
Abstract To reduce the clearance of sGLP-1, sGLP-1-HSA fusion protein was constructed by the fusion of the C-terminus of sGLP-1 to the N-terminus of HSA via an 6 amino acids linker and expressed in the pichia pastoris.Screen the strain with resistsnce of 4g/L G418. After fermentation in a 5L bioreactor,the expression of sGLP-1- HSA was induced by methanol.After 48h, the expression level reached 0.8g/L.The sGLP-1- HSA was abtained following a series of purification steps such as ultrafiltration, sephadex G-25 and blue- sepharose. The purity of the fermentation product could reach 95% by means of HPLC,and the total recovery yield could reach 60%.The biological activity of sGLP-1- HSA in GKⅡ diabetes rat and healthy rat suggested that the plasma glucose level was significantly lower after subcutaneous administration of sGLP-1- HSA..Expression of sGLP-1- HSA fusion protein can provide foudation for abtaining a larger quantity of recombinant sGLP-1- HSA for experimented and clinic studies.
|
Received: 19 February 2009
Published: 28 July 2009
|
|
|
|
[1] Vella A,Shah P, Basu R, et al. Effect of glucagons like peptide 1(736)amide on glucose effectiveness and insulin action people with type 2 diabetes .Diabetes,2000,49(4):611~617
[2] Wilding J P, Smith D M, Ghatei M A, et al. A role for glucagonslike peptide1 in the central regulation of feeding. Nature,1996,379:69~74
[3] Mentlein R, Gallwitz B, Schmidt W E, Dipeptidyl peptidase Ⅳ hydrolyses gastric inhibitory polypepetide, glucagonslike peptide1(736)amide, peptide histidine methionine and is responsible for their degradation in human serum. Eur J Biochen,1993,214:829~835
[4] Li Yuan, Shi Changhong, Lv Qiujun, et al.GLP1 Cterminal structures affect its blood glucose loweringfunction. J Pept Sci,2008,14:777~785
[5] Scorer C A, Clare J J, Mc Combie W R, et al. Rapid selection using G418 of high copy number transformants of Pichia pastoris for highlevel foreign gene expression. Bio/Technology,1994,12:181~184
[6] Clare J J , Rayment F B , Ballantine S P , et al . Highlevel expression of tetanus toxin fragment C in Pichia pastoris strains containing multiple tandem integrations of the gene . Bio/ Technology , 1991 ,9 (4) : 455 ~460
[7] 夏杰,徐殿胜,陆兵等.巴氏毕赤酵母SMD1168表达人溶菌酶的发酵条件.华东理工大学报,(自然科学版),2003,29(4):367~371
Xia J,Xu DS,Lu B,et al. Journal of East China University of Science and Technology(Natural Science Edition), 2003,29(4):367~371
[8] 刘彦丽,刘永东,王艳辉,等.毕赤酵母发酵生产重组人血清白蛋白高密度培养条件的研究.北京化工大学学报,2003,30(4):25~28
Liu Y L,Liu Y D,Wang Y H,et al. Journal of Beijing University of Chemical Technology (Natural Science Edition) 2003,30(4):25~28
[9] 邱荣德,李士云,陈俊刚,等.重组人血清白蛋白在Pichia pastoris 中的表达与纯化.生物化学与生物物理学报,2000, 32 (1) :59~62
Qiu R D,Li S Y,Chen J G,et al. Acta Biochimica et Biophysica Sinica,2000, 32 (1) :59~62 |
|
Viewed |
|
|
|
Full text
|
|
|
|
|
Abstract
|
|
|
|
|
Cited |
|
|
|
|
|
Shared |
|
|
|
|
|
Discussed |
|
|
|
|