Modification of the carboxy-terminal amino acid sequence alters the Escherichia coli expression of a gene encoding multiple repeats of a bovine growth hormone releasing factor analog

Trepod, Catherine M.; Mott, John E.

doi:10.1016/s0168-1656(00)00374-6

Cited by 3 publications

(2 citation statements)

References 34 publications

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“…1F) was then constructed and is designed to express a target protein as a C-terminal fusion to HN with an optional octapeptide K6EE C-terminus. Dipeptide glutamic acid (EE) or aspartic acid (DD) acidic residues at the C-terminus increased E. coli protein half-life (Parsell et al, 1990;Trepod and Mott, 2000). Consistent with this, the composite K6EE tag increased E. coli produced recombinant protein stability and yield (Williams, unpublished observations).…”

Section: Pvex Expression Vectorsmentioning

confidence: 77%

Thermostable tag (TST) protein expression system: Engineering thermotolerant recombinant proteins and vaccines

Luke

Carnes

Sun

et al. 2011

Journal of Biotechnology

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Methods to increase temperature stability of vaccines and adjuvants are needed to reduce dependence on cold chain storage. We report herein creation and application of pVEX expression vectors to improve vaccine and adjuvant manufacture and thermostability. Defined media fermentation yields of 6g/L thermostable toll-like receptor 5 agonist flagellin were obtained using an IPTG inducible pVEX-flagellin expression vector. Alternative pVEX vectors encoding Pyrococcus furiosus maltodextrin-binding protein (pfMBP) as a fusion partner improved Influenza hemagglutinin antigen vaccine solubility and thermostability. A pfMBP hemagglutinin HA2 domain fusion protein was a potent immunogen. Manufacturing processes that combined up to 5 g/L defined media fermentation yields with rapid, selective, thermostable pfMBP fusion protein purification were developed. The pVEX pfMBP-based thermostable tag (TST) platform is a generic protein engineering approach to enable high yield manufacture of thermostable recombinant protein vaccine components.

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Section: Pvex Expression Vectorsmentioning

confidence: 77%

Thermostable tag (TST) protein expression system: Engineering thermotolerant recombinant proteins and vaccines

Luke

Carnes

Sun

et al. 2011

Journal of Biotechnology

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“…Trepod C.M. et al [3] discovered that when the C-terminal amino acid residue sequence of bovine GHRH(1-30) was modulated, the E.coli expression of the gene encoding multiple repeats of the bovine GHRH(1-30) analog increased. Piquet G. et al [4] and Esposito P. et al [5] coupled the C-terminus of GHRH analog with polyethylene glycol (PEG).…”

Section: Introductionmentioning

confidence: 99%

A Novel hGHRH Analog and its Comparative Activity

Tang¹,

Du²,

Zhang³

et al. 2006

CCHTS

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Growth hormone releasing hormone is one of the hormones secreted from the hypothalamus. Because of its potential applications in agriculture and medicine, its short half-life and its expensive chemical synthesis, an analog with high GHRH activity and prolonged half-life was sought after. The fusion partner gene with 127 amino acid residues of the C-terminus from L-asparaginase was recombined with asp-pro-hGHRH(1-44) gene synthesized by PCR method to form one kind of fusion protein with unique acid labile linker Asp-Pro. The Pro-hGHRH(1-44) peptide was purified to homogeneity by means of cell disruption, washing, ethanol precipitation, acid hydrolysis, SP-Sephadex C-25, and Sephadex G-10 column chromatography. The peptide's molecular weight of 5,139 Da as measured by EIS-MS was coincident with the actual values. In the study of the activity, the doses of peptide were 0.1, 1.0, and 10 microg/ml for rat pituitary and 5 microg/ml for human pituitary. The peptide increased GH releases from rat pituitary in a concentration-dependent manner (P<0.05; P<0.01). At 1.0 microg/ml, there was a significant difference between Pro-Pro-hGHRH(1-44)-Gly-Gly-Cys and Pro-hGHRH(1-44) or Pro-Pro-hGHRH(1-44) (P<0.05), whereas the standard hGHRH(1-40) showed no measured rGH release. For human fetal pituitary, the Pro-hGHRH(1-44) peptides showed good GH-releasing activity, but there were no significant differences between them. The structure-activity relationship showed that for both rat and human fetal pituitary, the net GH-releasing activity of the Pro-hGHRH(1-44) analog was more than that of Pro-Pro-hGHRH(1-44). The results of the other hormones from human pituitary showed that the analog had good function-selectivity and species specificity.

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Structure and function relationships of three novel hGHRH-GGC analogs

Tang

Zhang

et al. 2010

Regulatory Peptides

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Modification of the carboxy-terminal amino acid sequence alters the Escherichia coli expression of a gene encoding multiple repeats of a bovine growth hormone releasing factor analog

Cited by 3 publications

References 34 publications

Thermostable tag (TST) protein expression system: Engineering thermotolerant recombinant proteins and vaccines

Thermostable tag (TST) protein expression system: Engineering thermotolerant recombinant proteins and vaccines

A Novel hGHRH Analog and its Comparative Activity

Structure and function relationships of three novel hGHRH-GGC analogs

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