Synthesis of phosphopeptides containing O-phosphoserine or O-phosphothreonine

Peptides containing phosphoserine or phosphothreonine were synthesized by solid phase methods. Phosphoserine and phosphothreonine were incorporated into peptides using Boc-diphenylphosphono esters of serine and threonine and standard DCC/HOBt coupling. The phenylphosphoesters were not removed when the peptides were cleaved from the resin by HF or by trifluoromethane sulfonic acid, but were subsequently removed by catalytic hydrogenation. Phosphopeptides were purified by HPLC and by Fe+3-Chelex chromatography and their identity verified by mass spectrometry. Two peptides, Leu-Arg-Arg-Ala-Ser(P)-Leu-Gly and Leu-Arg-Arg-Ala-Thr(P)-Leu-Gly, were prepared by both enzymatic and chemical methods and had identical properties.