Efficient introduction of alkene functionality into proteins in vivo.
The methionine analogue 2-amino-5-hexenoic acid (homoallylglycine, Hag) can be utilized by Escherichia coli in the initiation and elongation steps of protein biosynthesis. Use of an E. coli methionine auxotroph and Hag-supplemented medium resulted in replacement of ca. 85% of the methionine residues in mouse dihydrofolate reductase expressed under control of a bacteriophage T5 promoter. N-terminal sequencing indicated 92+/-5% occupancy of the initiator site by Hag. The vinyl function of Hag remains intact in the purified protein and suggests new chemistries for modification of natural and artificial proteins prepared in bacterial hosts.
van Hest
J C
JC
Department of Polymer Science and Engineering, University of Massachusetts, Amherst 01003, USA.
Tirrell
D A
DA
D000475:Alkenes / Q000378:metabolism*; D000818:Animals; D002208:Caproates / Q000378:metabolism*; D008715:Methionine / Q000378:metabolism; D051379:Mice; D014176:Protein Biosynthesis; D013762:Tetrahydrofolate Dehydrogenase / Q000096:biosynthesis* / Q000378:metabolism
C112998:2-amino-5-hexenoic acid; D000475:Alkenes; D002208:Caproates; D008715:Methionine; D013762:Tetrahydrofolate Dehydrogenase