title: A genetically encoded metabolically stable analogue of phosphotyrosine in Escherichia coli.
abstract: p-Carboxymethyl- l-phenylalanine (pCMF), a phosphotyrosine (pTyr) mimetic that is resistant to protein tyrosine phosphatase hydrolysis, was cotranslationally incorporated into proteins in Escherichia coli using an orthogonal amber suppressor tRNA/aminoacyl-tRNA synthetase (aaRS) pair. The pCMF-specific aaRS was identified from a large library of Methanococcus jannaschii tyrosyl-tRNA synthetase active-site mutants by a combination of positive and negative genetic selections. When pCMF was substituted for Tyr701 in human signal transducer and activator of transcription-1 (STAT1), a constitutively active mutant was obtained that dimerizes and binds a DNA oligonucleotide duplex that contains the M67 site recognized by Tyr701-phosphorylated STAT1. Genetic incorporation of pCMF into proteins should provide a new tool for the preparation of stable analogues of a wide array of phosphoproteins involved in signal transduction pathways, as well as the development of peptide-based, cellularly expressed inhibitors of pTyr binding proteins.
authors.forename: Jianming
authors.initials: J
authors.lastname: Xie
authors.forename: Lubica
authors.initials: L
authors.lastname: Supekova
authors.forename: Peter G
authors.initials: PG
authors.lastname: Schultz
mesh_terms: D019281:Dimerization; D004926:Escherichia coli / Q000235:genetics*; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000235:genetics / Q000378:metabolism*; D050794:STAT1 Transcription Factor / Q000378:metabolism; D015398:Signal Transduction
chemical_list: D050794:STAT1 Transcription Factor; D019000:Phosphotyrosine