Regioselective Carbon-Carbon Bond Formation in Proteins with Palladium Catalysis;New Protein Chemistry by Organometallic Chemistry

摘要

Palladium-catalyzed reactions have contributed to the advancement of many areas of organic chemistry,in particular,the synthesis of organic compounds such as natural products and polymeric materials.In this study,we have used a Mizoroki-Heck reaction for site-specific carbon-carbon bond formation in the Ras protein.This was performed by the following two steps;1)the His_6-fused Ras protein containing 4-iodo-L-phenylalanine at position 32(iF32-Ras-His)was prepared by genetic engineering and 2)the aryl iodide group on the iF32-Ras-His was coupled with vi-nylated biotin in the presence of a palladium catalyst.The biotin-ylation was confirmed by Western blotting and liquid chromatography-mass spectrometry(LC-MS).The regioselectivity of the Mizoroki-Heck reaction was furthermore confirmed by LC-MS/MS analysis.However,in addition to the biotinylated product(bF32-Ras-His),a dehalogenated product(F32-Ras-His)was detected by LC-MS/MS,This dehalogenation resulted from the undesired termination of the Mizoroki-Heck reaction due to steric and electrostatic hindrance around residue 32.The biotinylated Ras showed binding activity for the Ras-binding domain as its downstream target,Raf-1,with no sign of decomposition.This study is the first report of an application of Organometallic chemistry in protein chemistry.