Optimization of expression and purification of HSPA6 protein from Camelus dromedarius in E. coli
| Autor: | Abdulrahman M. Al-Senaidy, Mohammed S. Alanazi, Mohammad D. Bazzi, Wajahatullah Khan, Ajamaluddin Malik, Mohamed Elrobh |
|---|---|
| Jazyk: | angličtina |
| Předmět: |
0301 basic medicine
Fast protein liquid chromatography Size-exclusion chromatography lac operon medicine.disease_cause law.invention Hsp70 03 medical and health sciences law Native state medicine Incubation Escherichia coli Recombinant 030102 biochemistry & molecular biology Agricultural and Biological Sciences(all) Heat shock protein Chemistry Expression optimization 030104 developmental biology Biochemistry Yield (chemistry) Recombinant DNA Molecular chaperone Original Article General Agricultural and Biological Sciences |
| Zdroj: | Saudi Journal of Biological Sciences. (3):410-419 |
| ISSN: | 1319-562X |
| DOI: | 10.1016/j.sjbs.2015.04.017 |
| Popis: | The HSPA6, one of the members of large family of HSP70, is significantly up-regulated and has been targeted as a biomarker of cellular stress in several studies. Herein, conditions were optimized to increase the yield of recombinant camel HSPA6 protein in its native state, primarily focusing on the optimization of upstream processing parameters that lead to an increase in the specific as well as volumetric yield of the protein. The results showed that the production of cHSPA6 was increased proportionally with increased incubation temperature up to 37°C. Induction with 10μM IPTG was sufficient to induce the expression of cHSPA6 which was 100 times less than normally used IPTG concentration. Furthermore, the results indicate that induction during early to late exponential phase produced relatively high levels of cHSPA6 in soluble form. In addition, 5h of post-induction incubation was found to be optimal to produce folded cHSPA6 with higher specific and volumetric yield. Subsequently, highly pure and homogenous cHSPA6 preparation was obtained using metal affinity and size exclusion chromatography. Taken together, the results showed successful production of electrophoretically pure recombinant HSPA6 protein from Camelus dromedarius in Escherichia coli in milligram quantities from shake flask liquid culture. |
| Databáze: | OpenAIRE |
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