|
|
|
| The Expression and Purification of Human Heart-type Fatty Acid Binding Protein and Preparation of Lyophilized Protein |
| WU Jian-wei, CAI Lei, REN Yan-na, QIAN Wei, WANG Ji-hua, TANG Shi-xing |
| Guangzhou Wondfo Biotech Co.Ltd., Guangzhou 510663, China |
|
|
|
Abstract Objective: To obtain recombinant Human Heart-type Fatty Acid Binding Protein, detect its activity and prepare lyophilized protein. Methods: The CDs of Human H-FABP was searched from GenBank, synthesized into cloning vector. pET-28a(+)-H-FABP was constructed and transformed into E.coli BL21(DE3). After the optimal induction condition being researched, plenty of recombinant H-FABP was induced. Recombinant H-FABP was purified by Ni Sepharose 6 Fast Flow affinity chromatography and prepared into lyophilized protein. The activity of purified recombinant H-FABP and its lyophilized protein were detected by Wondfo H-FABP Test. Results: At the optimized induction condition(OD600/0.6, IPTG/0.4mmol/L, Induced for 4h at 30℃), recombinant H-FABP was expressed as soluble protein in E.coli BL21(DE3). High activity recombinant H-FABP was obtained with over 95% purity by Ni-chelating affinity chromatography. Lyophilized protein of recombinant H-FABP was stable for 12 days at 37℃ and 4 or more months at 25℃. Conclusion: The expression system established in this research is feasible and efficient. Lyophilized protein is stable enough to be provided as biological raw materials for further research.
|
|
Received: 24 December 2013
Published: 25 March 2014
|
|
|
|
|
| [1] 戚文航. 急性心肌梗死的流行病学及治疗进展. 现在实用医学, 2009, 16(111): 632-634. Qi W H. Epidemiology and treatment progress of acute myocardial infarction. Modern Practical Medicine, 2009, 16(111): 632-634.
[2] Wu H B, Panteghini M, Apple F, et al.Biochemical markers of cardiac damage: From traditional enzymes to cardiac-specific proteins.Scandinavian Journal of Clinical & Laboratory Investigation, 1999, 59(230):74-82.
[3] 潘柏申. 心脏标志物检测标准化研究进展. 中华检验医学杂志, 2007, 30(4):467-471. Pan B SH. Advance in detection standardization research of cardiac markers. Chin J Lab Med, 2007, 30(4): 467-471.
[4] Glatz J F, Vusse G J, Simoons M L, et al.Fatty acid-binding protein and the early detection of acute myocardial infarction.Clin Chim Acta, 1998, 272(1):87-92.
[5] Appie H, Kleine F, Glatz C, et al.Release of heart fatty acid-binding protein into plasma after acute myocardial infarction in man.Molecular and Cellular Biochemistry, 1992, 8(116):155-162.
[6] Nakata T, Hashimoto A, Hase M, et al.Human Heart-Type Fatty Acid-Binding Protein as an Early Diagnostic and Prognostic Marker in Acute Coronary Syndrome. Cardiology, 2003, 99(2):96-104.
[7] Junnichi I, Wang J H, Hiroyuki N, et al.Serum concentrations of myoglobin vs human heart-type cytoplasmic fatty acid-binding protein in early detection of acute myocardial infarction.Clinical Chemistry, 1997, 43(8): 1372-1378.
[8] Glatz J F, Vusse G J.Cellular fatty acid-binding protein: current concepts and future directions. Mol Cell Biochem, 1990, 6(89):237-251.
[9] Chmurzynska A.The multigene family of fatty acid-binding proteins (FABPs): Function, structure and polymorphism.Journal of Applied Genetics, 2006, 47(1):39-48.
[10] Schaap G, Spechtb Bernfried, Vussea J, et al.One-step purification of rat heart-type fatty acid-binding protein expressed in Escherichia coli.Journal of Chromatography B: Biomedical Sciences and Applications, 1996, 679(1-2): 61-67.
[11] 于志江, 陈旭等, 欧阳藩. 冷冻干燥技术在基因工程药物中的应用. 生物加工过程, 2005, 3(2):58-63. Yu ZH J, Chen X, Ouyang F. Freeze drying (lyophilization) of genetice engineering. Chinese Journal of Bioprocess Engineering, 2005, 31(2):58-63. |
|
Viewed |
|
|
|
Full text
|
|
|
|
|
Abstract
|
|
|
|
|
Cited |
|
|
|
|
| |
Shared |
|
|
|
|
| |
Discussed |
|
|
|
|
