mRNA secondary structure at start AUG codon is a key limiting factor for human protein expression in Escherichia coli

Weici Zhang, Weihua Xiao, Haiming Wei, Jian Zhang, Zhigang Tian

Research output: Contribution to journalArticle

39 Citations (Scopus)

Abstract

Codon usage and thermodynamic optimization of the 5′-end of mRNA have been applied to improve the efficiency of human protein production in Escherichia coli. However, high level expression of human protein in E. coli is still a challenge that virtually depends upon each individual target genes. Using human interleukin 10 (huIL-10) and interferon α (huIFN-α) coding sequences, we systematically analyzed the influence of several major factors on expression of human protein in E. coli. The results from huIL-10 and reinforced by huIFN-α showed that exposing AUG initiator codon from base-paired structure within mRNA itself significantly improved the translation of target protein, which resulted in a 10-fold higher protein expression than the wild-type genes. It was also noted that translation process was not affected by the retained short-range stem-loop structure at Shine-Dalgarno (SD) sequences. On the other hand, codon-optimized constructs of huIL-10 showed unimproved levels of protein expression, on the contrary, led to a remarkable RNA degradation. Our study demonstrates that exposure of AUG initiator codon from long-range intra-strand secondary structure at 5′-end of mRNA may be used as a general strategy for human protein production in E. coli.

Original languageEnglish (US)
Pages (from-to)69-78
Number of pages10
JournalBiochemical and Biophysical Research Communications
Volume349
Issue number1
DOIs
StatePublished - Oct 13 2006
Externally publishedYes

Fingerprint

Initiator Codon
Escherichia coli
Messenger RNA
Proteins
Interleukin-10
Escherichia coli Proteins
Codon
Genes
RNA Stability
Protein Biosynthesis
Thermodynamics
Interferons
RNA
Gene Expression
Degradation

Keywords

  • Codon optimization
  • E. coli
  • Interleukin 10
  • RNA stability
  • Secondary structure
  • Start codon

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

mRNA secondary structure at start AUG codon is a key limiting factor for human protein expression in Escherichia coli. / Zhang, Weici; Xiao, Weihua; Wei, Haiming; Zhang, Jian; Tian, Zhigang.

In: Biochemical and Biophysical Research Communications, Vol. 349, No. 1, 13.10.2006, p. 69-78.

Research output: Contribution to journalArticle

@article{2e048fe9a7384e9a8988861faaa6ce51,
title = "mRNA secondary structure at start AUG codon is a key limiting factor for human protein expression in Escherichia coli",
abstract = "Codon usage and thermodynamic optimization of the 5′-end of mRNA have been applied to improve the efficiency of human protein production in Escherichia coli. However, high level expression of human protein in E. coli is still a challenge that virtually depends upon each individual target genes. Using human interleukin 10 (huIL-10) and interferon α (huIFN-α) coding sequences, we systematically analyzed the influence of several major factors on expression of human protein in E. coli. The results from huIL-10 and reinforced by huIFN-α showed that exposing AUG initiator codon from base-paired structure within mRNA itself significantly improved the translation of target protein, which resulted in a 10-fold higher protein expression than the wild-type genes. It was also noted that translation process was not affected by the retained short-range stem-loop structure at Shine-Dalgarno (SD) sequences. On the other hand, codon-optimized constructs of huIL-10 showed unimproved levels of protein expression, on the contrary, led to a remarkable RNA degradation. Our study demonstrates that exposure of AUG initiator codon from long-range intra-strand secondary structure at 5′-end of mRNA may be used as a general strategy for human protein production in E. coli.",
keywords = "Codon optimization, E. coli, Interleukin 10, RNA stability, Secondary structure, Start codon",
author = "Weici Zhang and Weihua Xiao and Haiming Wei and Jian Zhang and Zhigang Tian",
year = "2006",
month = "10",
day = "13",
doi = "10.1016/j.bbrc.2006.07.209",
language = "English (US)",
volume = "349",
pages = "69--78",
journal = "Biochemical and Biophysical Research Communications",
issn = "0006-291X",
publisher = "Academic Press Inc.",
number = "1",

}

TY - JOUR

T1 - mRNA secondary structure at start AUG codon is a key limiting factor for human protein expression in Escherichia coli

AU - Zhang, Weici

AU - Xiao, Weihua

AU - Wei, Haiming

AU - Zhang, Jian

AU - Tian, Zhigang

PY - 2006/10/13

Y1 - 2006/10/13

N2 - Codon usage and thermodynamic optimization of the 5′-end of mRNA have been applied to improve the efficiency of human protein production in Escherichia coli. However, high level expression of human protein in E. coli is still a challenge that virtually depends upon each individual target genes. Using human interleukin 10 (huIL-10) and interferon α (huIFN-α) coding sequences, we systematically analyzed the influence of several major factors on expression of human protein in E. coli. The results from huIL-10 and reinforced by huIFN-α showed that exposing AUG initiator codon from base-paired structure within mRNA itself significantly improved the translation of target protein, which resulted in a 10-fold higher protein expression than the wild-type genes. It was also noted that translation process was not affected by the retained short-range stem-loop structure at Shine-Dalgarno (SD) sequences. On the other hand, codon-optimized constructs of huIL-10 showed unimproved levels of protein expression, on the contrary, led to a remarkable RNA degradation. Our study demonstrates that exposure of AUG initiator codon from long-range intra-strand secondary structure at 5′-end of mRNA may be used as a general strategy for human protein production in E. coli.

AB - Codon usage and thermodynamic optimization of the 5′-end of mRNA have been applied to improve the efficiency of human protein production in Escherichia coli. However, high level expression of human protein in E. coli is still a challenge that virtually depends upon each individual target genes. Using human interleukin 10 (huIL-10) and interferon α (huIFN-α) coding sequences, we systematically analyzed the influence of several major factors on expression of human protein in E. coli. The results from huIL-10 and reinforced by huIFN-α showed that exposing AUG initiator codon from base-paired structure within mRNA itself significantly improved the translation of target protein, which resulted in a 10-fold higher protein expression than the wild-type genes. It was also noted that translation process was not affected by the retained short-range stem-loop structure at Shine-Dalgarno (SD) sequences. On the other hand, codon-optimized constructs of huIL-10 showed unimproved levels of protein expression, on the contrary, led to a remarkable RNA degradation. Our study demonstrates that exposure of AUG initiator codon from long-range intra-strand secondary structure at 5′-end of mRNA may be used as a general strategy for human protein production in E. coli.

KW - Codon optimization

KW - E. coli

KW - Interleukin 10

KW - RNA stability

KW - Secondary structure

KW - Start codon

UR - http://www.scopus.com/inward/record.url?scp=33748316617&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=33748316617&partnerID=8YFLogxK

U2 - 10.1016/j.bbrc.2006.07.209

DO - 10.1016/j.bbrc.2006.07.209

M3 - Article

C2 - 16930549

AN - SCOPUS:33748316617

VL - 349

SP - 69

EP - 78

JO - Biochemical and Biophysical Research Communications

JF - Biochemical and Biophysical Research Communications

SN - 0006-291X

IS - 1

ER -