Ultrafast Dynamics Study On Energy Transfer Between Tryptophan And Tyrosine: From Peptides To Proteins

Proteins are fundamental to the living body.Tryptophan and tyrosine are crucial intrinsic fluorescent amino acids that participate in various complex mechanisms and exert biological functions of proteins.Investigating the interactions between those amino acids can provide us insights into the scientific problems related to protein structure and function.Such research plays a critical role in advancing protein science and life sciences.Therefore,we have employed a bottom-up research approach to systematically explore the interactions between tryptophan and tyrosine.Our studies do not only uncover their diverse properties but also give an essential theoretical and experimental groundwork for further exploration of protein structure and function.Firstly,this thesis investigated the fluorescence kinetic characteristics of tryptophan and tyrosine in water / n-propanol mixed solvents and deep eutectic solvents.By comparing the fluorescence properties of tryptophan and peptides in water / npropanol mixed solvents,we explored how tryptophan responses to the solvent properties and peptide structural changes.We also observed two solvation processes and two “isomers” conformations of tryptophan in deep eutectic solvents,which we attributed to the special solvent environment and rich hydrogen bond network of deep eutectic solvents.Furthermore,we explored the fluorescence kinetic characteristics of tyrosine and tyrosinate.Our results provided an important theoretical support for subsequent interaction studies between tryptophan and tyrosine.Secondly,we designed and synthesized a series of model peptides,WPn Y(n = 1,2,3,5,8),to provide different distances between tryptophan and tyrosine.We conducted a comprehensive analysis of F?rster resonance energy transfer(FRET)between tryptophan and tyrosine in the model peptides through both experimental and theoretical approaches.Our results demonstrated that ultrafast FRET between tryptophan and tyrosine could alter the intrinsic fluorescence dynamics of tryptophan and significantly affected the analysis of protein hydration.Moreover,we proposed a hybrid lifetime separation scheme that we successfully applied to extract the FRET characteristic lifetime in the model peptides.This scheme provided an important solution for further studies of intra protein amino acid interactions.Thirdly,we studied the electron transfer properties in the model peptides.Our results indicated that the conversion process between tryptophan radicals and photoproducts in the peptides is limited to varying degrees,which greatly reduced the yield of tryptophan triplet state.We found that a “dyads” structure was formed between tryptophan and tyrosine in WPY,and the content of free radicals and triplets was reduced by proton coupled electron transfer(PCET)in direct formula.In other peptides,the distance between tryptophan and tyrosine were greater,so only indirect PCET could occur.Both modes of PCET could significantly reduce the content of tryptophan radicals and triplet states,providing important experimental evidence for further understanding the self-protection mechanism of clearing radicals in proteins.Finally,we extended the FRET dynamics study between tryptophan and tyrosine from model peptides to the protein monellin.Our results demonstrated that FRET between tryptophan and tyrosine was significantly affected by protein hydration,confirming the conclusions from the model peptide studies.We proposed a simple scheme that could be used to judge whether protein hydration was affected by the FRET effect.Additionally,we successfully separated the characteristic lifetime of FRET in proteins,and discussed the feasibility of this characteristic lifetime for fine structural analysis of proteins.Our findings suggested that this characteristic lifetime could serve as a new high-sensitivity natural “spectral ruler”.In summary,this thesis elucidated in detail the energy transfer and electron transfer mechanisms between tryptophan and tyrosine in peptides and proteins.Our study provided a comprehensive and detailed theoretical foundation for evaluating protein hydration,developing protein intrinsic fluorescence “spectral ruler”,and understanding the self-protection mechanisms of proteins.