Overexpression in Escherichia coli, purification and characterization of Thermoanaerobacter tengcongensis elongation factor G with multiple rare codons

Liqiang Zhang; Peng Xue; Hongjie Zhang

doi:10.2174/092986607781483732

Overexpression in Escherichia coli, purification and characterization of Thermoanaerobacter tengcongensis elongation factor G with multiple rare codons

Liqiang Zhang, Peng Xue, Hongjie Zhang

Research output: Contribution to journal › Article › peer-review

3 Scopus citations

Abstract

The fusA gene encoding a thermophilic protein EF-G with multiple rare condons was cloned from Thermoanaerobacter tengcongensis (TteEF-G) and overexpressed in Escherichia coli by cotransfering a RIG plasmid to overcome the potential codon-bias problem originated from Arg, Ile and Gly. The recombinant protein was identified by MALDITOF-MS for molecular mass with approximation of 76 kDa and by trypsin digestion coupled LC-MS/MS for peptide sequence coverage of 61.3%. The in vivo complementary assay indicates that TteEF-G could significantly rescue the E. coli LJ14 (frr^ts) at the non-permission temperature of 42γC in the bi-transformant of TteRRF and TteEF-G. This study indicated that coexpression of rare codons' cognate tRNA is a useful method for protein overexpression in E. coli.

Original language	English (US)
Pages (from-to)	804-810
Number of pages	7
Journal	Protein and Peptide Letters
Volume	14
Issue number	8
DOIs	https://doi.org/10.2174/092986607781483732
State	Published - Aug 2007
Externally published	Yes

Keywords

Elongation factor G
Gene expression
Purification and characterization
RIG plasmid
Thermoanaerobacter tengcongensis

ASJC Scopus subject areas

Structural Biology
Biochemistry

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10.2174/092986607781483732

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title = "Overexpression in Escherichia coli, purification and characterization of Thermoanaerobacter tengcongensis elongation factor G with multiple rare codons",

abstract = "The fusA gene encoding a thermophilic protein EF-G with multiple rare condons was cloned from Thermoanaerobacter tengcongensis (TteEF-G) and overexpressed in Escherichia coli by cotransfering a RIG plasmid to overcome the potential codon-bias problem originated from Arg, Ile and Gly. The recombinant protein was identified by MALDITOF-MS for molecular mass with approximation of 76 kDa and by trypsin digestion coupled LC-MS/MS for peptide sequence coverage of 61.3%. The in vivo complementary assay indicates that TteEF-G could significantly rescue the E. coli LJ14 (frrts) at the non-permission temperature of 42γC in the bi-transformant of TteRRF and TteEF-G. This study indicated that coexpression of rare codons' cognate tRNA is a useful method for protein overexpression in E. coli.",

keywords = "Elongation factor G, Gene expression, Purification and characterization, RIG plasmid, Thermoanaerobacter tengcongensis",

author = "Liqiang Zhang and Peng Xue and Hongjie Zhang",

year = "2007",

month = aug,

doi = "10.2174/092986607781483732",

language = "English (US)",

volume = "14",

pages = "804--810",

journal = "Protein and Peptide Letters",

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T1 - Overexpression in Escherichia coli, purification and characterization of Thermoanaerobacter tengcongensis elongation factor G with multiple rare codons

AU - Zhang, Liqiang

AU - Xue, Peng

AU - Zhang, Hongjie

PY - 2007/8

Y1 - 2007/8

N2 - The fusA gene encoding a thermophilic protein EF-G with multiple rare condons was cloned from Thermoanaerobacter tengcongensis (TteEF-G) and overexpressed in Escherichia coli by cotransfering a RIG plasmid to overcome the potential codon-bias problem originated from Arg, Ile and Gly. The recombinant protein was identified by MALDITOF-MS for molecular mass with approximation of 76 kDa and by trypsin digestion coupled LC-MS/MS for peptide sequence coverage of 61.3%. The in vivo complementary assay indicates that TteEF-G could significantly rescue the E. coli LJ14 (frrts) at the non-permission temperature of 42γC in the bi-transformant of TteRRF and TteEF-G. This study indicated that coexpression of rare codons' cognate tRNA is a useful method for protein overexpression in E. coli.

AB - The fusA gene encoding a thermophilic protein EF-G with multiple rare condons was cloned from Thermoanaerobacter tengcongensis (TteEF-G) and overexpressed in Escherichia coli by cotransfering a RIG plasmid to overcome the potential codon-bias problem originated from Arg, Ile and Gly. The recombinant protein was identified by MALDITOF-MS for molecular mass with approximation of 76 kDa and by trypsin digestion coupled LC-MS/MS for peptide sequence coverage of 61.3%. The in vivo complementary assay indicates that TteEF-G could significantly rescue the E. coli LJ14 (frrts) at the non-permission temperature of 42γC in the bi-transformant of TteRRF and TteEF-G. This study indicated that coexpression of rare codons' cognate tRNA is a useful method for protein overexpression in E. coli.

KW - Elongation factor G

KW - Gene expression

KW - Purification and characterization

KW - RIG plasmid

KW - Thermoanaerobacter tengcongensis

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JO - Protein and Peptide Letters

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Overexpression in Escherichia coli, purification and characterization of Thermoanaerobacter tengcongensis elongation factor G with multiple rare codons

Abstract

Keywords

ASJC Scopus subject areas

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