Design, total chemical synthesis, and binding properties of a [Leu-91-N1-methyl-7-azaTrp]Ras-binding domain of c-Raf-1
AUTOR(ES)
Sydor, Jens R.
FONTE
The National Academy of Sciences
RESUMO
The Ras-binding domain (RBD) of c-Raf-1 has been synthesized chemically, taking advantage of the chemical ligation of two peptide fragments of the protein. This procedure allowed incorporation of an unnatural amino acid (N1-methyl-7-azatryptophan) at position 91 of RBD, producing a protein with fluorescent properties distinct from and distinguishable from those of proteins containing the natural fluorophore tryptophan. The resulting protein was shown to interact with Ras in a manner that was almost indistinguishable from that of unmodified RBD based on transient kinetic monitoring of the binding event. Modified RBD containing the l-isomer of the unnatural amino acid or its racemic d,l mixture appeared to interact identically with Ras. The approach demonstrates a general procedure for the introduction of unnatural amino acids that can be used for monitoring protein–protein interactions and for the introduction of an unnatural backbone structure at strategic positions.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=22153Documentos Relacionados
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