Studies On The Effect Of Phosphorylation On The Peptidyl-prolyl Imide Bond Cis/trans Isomerization

Proline plays an unique and key role in peptide and protein structure. Thecis/trans isomerization of peptidyl-prolyl imide bond formed by proline and theamino acids preceding proline acts as a molecular switch in controlling many lifeprocess in vivo. Among all the factors influencing the cis/trans isomerization, theside-chain O-phosphorylation on the amino acids preceding proline, a well-knownreversible posttranslational modification itself, is important as it might exert directeffects as well as indirect effects on this kind of isomerization. Studies in this fieldmight further illuminate how the phosphorylation regulates the protein function.Two sorts of tetrapeptide models were designed to explore the effects ofphosphorylation on peptidyl-prolyl imide bond cis/trans isomeriztion in this thesis.Phosphopeptides and unphosphopeptides with Ser/Thr/Tyr -Pro motif weresynthesized by global and step-wise solid phase synthesis method based on Fmocstrategy. A new quantitative analysis protocol for the cis content of those peptideswas revealed during the HPLC separation. The MS data for protected amino acidsindicated a new rearrangement reaction via five-membered ring intermediate in ESIfragments. And the fragmentation differences between the ESI-MS andMALDI-TOF-MS were also discussed.Studies on the cis content of all these model compounds were carried out by1H-NMR method. Based on these data, the effects of side-chain, phophorylationprocess and charges were investigated on the cis/trans isomerization of thephosphorylated peptidyl-prolyl amide bond. The possible mechanism were proposed,in the aspects of the conformation, structure and molecular interaction, about theinfluence of phsophorylation on peptidyl-prolyl imide bond cis/transisomerization through 2D NMR and CD studies. Thereafter, the molecularmodeling studies were performed on these model peptides in an attempt to furtherestablish the effect of phsophorylation on the peptidyl-prolyl imide bondcis/trans isomerization and the results were basically consistent with theexperimental data.According to the research, it was concluded that the phosphorylation itself thatcooperated with the side-chain and charge effects influcened the peptidyl-prolylimide bond cis/trans isomerization and subsequently acted as a molecular switch tocontrol the physiological process in life.