A new strategy for glycoprotein synthesis: ligation of synthetic glycopeptides with truncated proteins expressed in E. coli as TEV protease cleavable fusion protein

[Display omitted] We report here the use of TEV protease cleavable fusion proteins to produce glycosylated bioactive peptides and proteins. Bacterial expression was utilized to produce two fusion proteins, GPRT-C37-H6 and His-tagged interleukin-2 (amino acids 6–133), which when cleaved by the tobacc...

Full description

Saved in:
Bibliographic Details
Published in:Bioorganic & medicinal chemistry 2005-02, Vol.13 (3), p.909-915
Main Authors: Tolbert, Thomas J., Franke, Dirk, Wong, Chi-Huey
Format: Article
Language:English
Subjects:
Base Sequence
Bioconversions. Hemisynthesis
Biological and medical sciences
Biotechnology
Cloning, Molecular
DNA Primers
Electrophoresis, Polyacrylamide Gel
Endopeptidases - biosynthesis
Escherichia coli - metabolism
Fundamental and applied biological sciences. Psychology
Glycopeptides - chemistry
Glycoproteins - biosynthesis
Glycoproteins - chemistry
Glycoproteins - genetics
Glycoproteins - isolation & purification
Humans
Methods. Procedures. Technologies
Recombinant Fusion Proteins - biosynthesis
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:[Display omitted] We report here the use of TEV protease cleavable fusion proteins to produce glycosylated bioactive peptides and proteins. Bacterial expression was utilized to produce two fusion proteins, GPRT-C37-H6 and His-tagged interleukin-2 (amino acids 6–133), which when cleaved by the tobacco etch virus NIa protease (TEV protease) to generate HIV entry inhibitor peptide C37-H6 and a truncated version of the cytokine interleukin-2, both containing N-terminal cysteines. The N-terminal cysteine containing C37-H6 and truncated interleukin-2 were then joined to a synthetic glycopeptide thioester utilizing native chemical ligation under nondenaturing and denaturing conditions, respectively. The ligations of the glycopeptide to the C37-H6 peptide and the truncated interleukin-2 protein both proceeded in high yield, though the size, and physical properties of the two polypeptides differ greatly.
ISSN:0968-0896
1464-3391
DOI:10.1016/j.bmc.2004.06.047