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Partial purification of human parathyroid hormone 1-84 as a thioredoxin fusion form in recombinant Escherichia coli by thermoosmotic shock.
- Source :
-
Protein expression and purification [Protein Expr Purif] 2006 Sep; Vol. 49 (1), pp. 32-8. Date of Electronic Publication: 2006 Mar 29. - Publication Year :
- 2006
-
Abstract
- A modified purification method, thermoosmotic shock (osmotic shock coupled with heat-treatment) for heat-stable proteins, was devised in the purification of Trx-hPTH (1-84) (human parathyroid hormone coupled with thioredoxin as a fusion partner) from E. coli. Thermoosmotic shock can integrate the functions of extraction and crude separation of fusion protein Trx-hPTH (1-84). To improve the purification efficiency, thermoosmotic shock conditions were optimized and achieved as follows: the optimized high osmotic solution containing 20mM Tris-HCl buffer (pH 8.0), 1mM EDTA, and 25% sucrose; the low osmotic solution containing 20mM Tris-HCl buffer (pH 8.0), 1mM EDTA, and the heat-treatment temperature of 100 degrees C for 10 min. Using this method, the purity of Trx-hPTH (1-84) was up to 73% and the yield was up to 72%, respectively. In addition, the two separation methods of both thermoosmotic shock and affinity chromatography have been compared, indicating that thermoosmotic shock is an economical and feasible way for the fusion protein separation. Besides, the thermoosmotic shock method may be used for the purification of some proteins of thermal stability without N-terminal His-tag.
- Subjects :
- Chromatography, Affinity
Cyclic AMP metabolism
Edetic Acid
Escherichia coli genetics
Humans
Osmotic Pressure
Parathyroid Hormone genetics
Parathyroid Hormone pharmacology
Recombinant Fusion Proteins genetics
Recombinant Fusion Proteins pharmacology
Temperature
Thioredoxins genetics
Escherichia coli metabolism
Parathyroid Hormone isolation & purification
Parathyroid Hormone metabolism
Recombinant Fusion Proteins isolation & purification
Recombinant Fusion Proteins metabolism
Thioredoxins isolation & purification
Thioredoxins metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 1046-5928
- Volume :
- 49
- Issue :
- 1
- Database :
- MEDLINE
- Journal :
- Protein expression and purification
- Publication Type :
- Academic Journal
- Accession number :
- 16624572
- Full Text :
- https://doi.org/10.1016/j.pep.2006.03.004