Site-specific incorporation of the mucin-type N-acetylgalactosamine-alpha-O-threonine into protein in Escherichia coli.
Glycosylation is a prevalent posttranslational process capable of augmenting and modulating protein function. Efficient synthesis of high-purity, homogeneous glycoproteins is essential for the study of unique protein glycoforms and for the manufacture of therapeutically relevant forms. A promising new strategy for controlled in vivo synthesis of glycoproteins was recently established using suppressor tRNA technology. Using an evolved tRNA aminoacyl synthetase-tRNA pair from Methanococcus jannaschii, the glycosyl amino acid, N-acetylglucosamine-beta-O-serine (GlcNAc-beta-Ser), was site-specifically introduced into proteins cotranslationally in Escherichia coli. Herein, we report the evolution of a new tRNA aminoacyl synthetase-tRNA pair that has expanded the repertoire of glycoproteins that can be expressed in E. coli to contain the other major O-linked glycan, N-acetylgalactosamine-alpha-O-threonine (GalNAc-a-Thr).
Xu
Ran
R
Department of Chemistry, and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 Torrey Pines Road, La Jolla, California 92037, USA.
Hanson
Sarah R
SR
Zhang
Zhiwen
Z
Yang
Yu-Ying
YY
Schultz
Peter G
PG
Wong
Chi-Huey
CH
D000116:Acetylgalactosamine / Q000031:analogs & derivatives* / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000096:biosynthesis / Q000235:genetics; D009077:Mucins / Q000235:genetics / Q000378:metabolism*; D009211:Myoglobin / Q000096:biosynthesis / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D021241:Spectrometry, Mass, Electrospray Ionization; D013912:Threonine / Q000031:analogs & derivatives* / Q000235:genetics / Q000378:metabolism*; D014448:Tyrosine-tRNA Ligase / Q000235:genetics / Q000378:metabolism
D029968:Escherichia coli Proteins; D009077:Mucins; D009211:Myoglobin; D011994:Recombinant Proteins; D013912:Threonine; D014448:Tyrosine-tRNA Ligase; D000116:Acetylgalactosamine