Hunting Phosphorylation

A straightforward method for severing large peptides precisely at phosphorylated residues may make it easier to pinpoint phosphorylation sites in proteins (J. Am. Chem. Soc., DOI: 10.1021/ja8023719). Cells use phosphorylation as a signaling mechanism. To understand what these chemical flags mean, scientists must first be able to find them. Ryan R. Julian and Jolene K. Diedrich at the University of California, Riverside, devised a photofragmentation approach that permits them to selectively break peptide backbones at phosphorylated serine and threonine residues. They used a strong base (barium hydroxide) to eliminate the phosphate group and then added a thiol-containing naphthyl chromophore (shown). The resulting carbon-sulfur bond (red) cleaves homolytically when the chromophore absorbs an ultraviolet photon of the appropriate wavelength. The carbon radical product rearranges to cleave the peptide backbone into fragments that can be analyzed by mass spectrometry to determine the original phosphorylation sites.