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PMID: 17126029 Published · ppublish English Journal Article Research Support, N.I.H., Extramural Research Support, U.S. Gov't, Non-P.H.S.

Effect of osmotic stress and heat shock in recombinant protein overexpression and crystallization.

Protein expression and purification ·Vol. 52 ·No. 2 ·2007-04-00 ·Pages 280-5

Oganesyan N, Ankoudinova I, Kim SH, Kim R

Abstract

Overexpressed recombinant proteins in bacteria often tend to misfold and accumulate as soluble aggregates and/or inclusion bodies. A strategy for improving the level of expression of recombinant proteins in a soluble native form is to increase the cellular concentration of osmolytes or of chaperones. This can be accomplished by growing the bacterial cells in the presence of high salt, sorbitol, and betaine as well as exposing the cells to a heat shock step. Our results suggest that by growing the cells under varied conditions one may be able to express targets as soluble proteins (from previously insoluble targets) and to improve the chances of their crystallization.

MeSH Terms
Crystallization Escherichia coli/genetics Gene Expression Heat-Shock Response Recombinant Proteins/chemistry,genetics,metabolism
Chemicals
Recombinant Proteins
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Oganesyan Natalia
Bio-Rad Laboratories, Life Science Group, 2000 Alfred Nobel Drive, Hercules, CA 94547, USA.
Ankoudinova Irina
Kim Sung-Hou
Kim Rosalind
References (28)
28 references, click to expand
  1. Increase of soluble expression in Escherichia coli cytoplasm by a protein disulfide isomerase gene fusion system.
    Protein Expr Purif. 2005 Dec;44(2):155-61 PMID: 15882951
  2. Ligation-independent cloning of PCR products (LIC-PCR).
    Nucleic Acids Res. 1990 Oct 25;18(20):6069-74 PMID: 2235490
  3. Expression of recombinant zeta-crystallin in Escherichia coli with the help of GroEL/ES and its purification.
    Protein Expr Purif. 2001 Mar;21(2):260-7 PMID: 11237687
  4. Production of Lewis x tetrasaccharides by metabolically engineered Escherichia coli.
    Chembiochem. 2006 Feb;7(2):359-65 PMID: 16381046
  5. Tight binding of deoxyribonucleotide triphosphates to human thymidine kinase 2 expressed in Escherichia coli. Purification and partial characterization of its dimeric and tetrameric forms.
    Biochemistry. 2003 Dec 30;42(51):15158-69 PMID: 14690426
  6. The osmophobic effect: natural selection of a thermodynamic force in protein folding.
    J Mol Biol. 2001 Jul 27;310(5):955-63 PMID: 11502004
  7. Protein misfolding and inclusion body formation in recombinant Escherichia coli cells overexpressing Heat-shock proteins.
    J Biol Chem. 1996 May 10;271(19):11141-7 PMID: 8626659
  8. DnaK and DnaJ facilitated the folding process and reduced inclusion body formation of magnesium transporter CorA overexpressed in Escherichia coli.
    Protein Expr Purif. 2003 Dec;32(2):221-31 PMID: 14965767
  9. Native folding of aggregation-prone recombinant proteins in Escherichia coli by osmolytes, plasmid- or benzyl alcohol-overexpressed molecular chaperones.
    Cell Stress Chaperones. 2005 Winter;10(4):329-39 PMID: 16333986
  10. The Hsp70 and Hsp60 chaperone machines.
    Cell. 1998 Feb 6;92(3):351-66 PMID: 9476895
  11. Enhancement of the solubility of proteins overexpressed in Escherichia coli by heat shock.
    J Mol Microbiol Biotechnol. 2002 Nov;4(6):519-24 PMID: 12432951
  12. Protein production by auto-induction in high density shaking cultures.
    Protein Expr Purif. 2005 May;41(1):207-34 PMID: 15915565
  13. Proline inhibits aggregation during protein refolding.
    Protein Sci. 2000 Feb;9(2):344-52 PMID: 10716186
  14. A novel strategy for production of a highly expressed recombinant protein in an active form.
    FEBS Lett. 1991 Dec 16;295(1-3):10-2 PMID: 1765138
  15. Uptake and synthesis of compatible solutes as microbial stress responses to high-osmolality environments.
    Arch Microbiol. 1998 Oct;170(5):319-30 PMID: 9818351
  16. Preparation of selenomethionyl proteins for phase determination.
    Methods Enzymol. 1997;276:523-30 PMID: 9048379
  17. Protein synthesis inhibitors and ethanol selectively enhance heterologous expression of P450s and related proteins in Escherichia coli.
    Arch Biochem Biophys. 1999 Jul 1;367(1):129-36 PMID: 10375408
  18. Manipulating the amyloid-beta aggregation pathway with chemical chaperones.
    J Biol Chem. 1999 Nov 12;274(46):32970-4 PMID: 10551864
  19. Soluble expression of recombinant proteins in the cytoplasm of Escherichia coli.
    Microb Cell Fact. 2005 Jan 04;4(1):1 PMID: 15629064
  20. Crystal structure of the conserved hypothetical protein MPN330 (GI: 1674200) from Mycoplasma pneumoniae.
    Proteins. 2005 Feb 1;58(2):504-8 PMID: 15562512
  21. Chemical chaperones regulate molecular chaperones in vitro and in cells under combined salt and heat stresses.
    J Biol Chem. 2001 Oct 26;276(43):39586-91 PMID: 11517217
  22. Expression and purification of histidine-tagged bacteriophage T7 DNA polymerase.
    Protein Expr Purif. 2005 Feb;39(2):247-53 PMID: 15642476
  23. Chaperonin-assisted folding of glutamine synthetase under nonpermissive conditions: off-pathway aggregation propensity does not determine the co-chaperonin requirement.
    Protein Sci. 2000 Dec;9(12):2405-12 PMID: 11206062
  24. A new strategy for the synthesis of glycoproteins.
    Science. 2004 Jan 16;303(5656):371-3 PMID: 14726590
  25. Overexpression of DsbC and DsbG markedly improves soluble and functional expression of single-chain Fv antibodies in Escherichia coli.
    Protein Expr Purif. 2002 Nov;26(2):218-28 PMID: 12406675
  26. Mutational and structural-based analyses of the osmolyte effect on protein stability.
    J Biochem. 2004 Jun;135(6):701-8 PMID: 15213245
  27. Recombinant protein expression in Escherichia coli.
    Curr Opin Biotechnol. 1999 Oct;10(5):411-21 PMID: 10508629
  28. Recombinant protein folding and misfolding in Escherichia coli.
    Nat Biotechnol. 2004 Nov;22(11):1399-408 PMID: 15529165
Article Info
Journal
Protein expression and purification
Abbr.
Protein Expr Purif
ISSN
1046-5928
DOI
10.1016/j.pep.2006.09.015
Published
2007-04-00
Epub
2006-00-10
Pages
280-5
Language
English
Region
United States
NLM ID
9101496
PMCID
PMC1865119
Subset
IM
Grants
NIGMS NIH HHS · P50 GM062412 · United States
NIGMS NIH HHS · GM 62412 · United States
NIGMS NIH HHS · P50 GM062412-050002 · United States
External Links
PubMed source DOI: 10.1016/j.pep.2006.09.015 PMC Full Text View journal details
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