Site-specific incorporation of glycosylated serine and tyrosine derivatives into proteins.
Glycosylation of proteins can have a dramatic effect on their physical, chemical, and biological properties. Analogues of dihydrofolate reductase and firefly luciferase containing glycosylated amino acids at single, predetermined sites have been elaborated. Misacylated suppressor tRNAs activated with glycosylated serine and tyrosine derivatives were used for suppression of the nonsense codons in a cell-free protein biosynthesizing system, thereby permitting the preparation of the desired glycosylated proteins. In this fashion, it was possible to obtain proteins containing both mono- and diglycosylated amino acids, including glycosylated serine and tyrosine moieties. For the modified firefly luciferases, the effect of these substitutions on the wavelength of the light emitted by firefly luciferase was investigated. The maximum wavelength for mutants containing peracetylated glycosylated serine derivatives at position 284 showed a red shift in the emission spectra. For mutants containing glycosylated tyrosines, the red shift was observed only when the carbohydrate moiety was fully deacetylated.
Fahmi
Nour Eddine
NE
Department of Chemistry, University of Virginia, Charlottesville, Virginia 22901, USA.
Dedkova
Larisa
L
Wang
Bixun
B
Golovine
Serguei
S
Hecht
Sidney M
SM
D000818:Animals; D004926:Escherichia coli; D004952:Esters / Q000737:chemistry; D048888:Fireflies; D006031:Glycosylation; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism*; D012343:RNA, Transfer / Q000737:chemistry; D012694:Serine / Q000378:metabolism*; D013762:Tetrahydrofolate Dehydrogenase / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism*
D004952:Esters; D011506:Proteins; D014443:Tyrosine; D012694:Serine; D012343:RNA, Transfer; D013762:Tetrahydrofolate Dehydrogenase