Characterizing The Structures And Spectra Of Chromophores In Condensed Phase By Means Of Multiscale Models

The green fluorescent proteins(GFP)is one of an important class of proteins that ex-hibit strong visible fluorescence without the requirement of cofactors or other enzymes.In the last two decades,GFP and its mutants have changed from a nearly unknown protein to be the one widely applied in many fields.To regulate spectral properties,such as the bright-ness and the emission maxima,many GFP mutants have been devised.In general,there are two types of mutation.The first type occurs mutations on the amino acid residues that participate in the formation of the chromophore,and the other type occurs on the amino acid residues that contact the chromophore in its close vicinity.The changes of spectra in the first mutant type have been attributed to changes in the chromophore structure with modulations on the conjugation length.The representative examples of the first mutant type are the blue fluorescent protein(BFP)and the cyan fluorescent protein(CFP).Although red fluorescent protein(RFP)is not directly derived from the aequorea victoria GFP,it can also be seen as the first mutant type.In the second mutant type,changes of spectra originate from the molecular environment.The yellow fluorescent protein(YFP)is a representative example of the second mutant type.The replacement of Thr203 with other aromatic amino acids(His,Trp,Phe,and Tyr)leads to the longest wavelength emission of all GFPs with different protein environment.To design GFP variants with improved wavelength and brightness in living cells,it is essential to theoretically calculate optical properties of the chromophores,understand the structure-property relationship,and reveal the microscopic environmental effect on the chromophore's photophysical and photochemical processes.Although GFP and its mutants have been extensively investigated by many experimental and theoretical works,it is still unclear on the microscopic origin of the photophysical and photochemical processes taken place in those proteins,the geometrical and electronic structures of the concerned states related to these processes,the large variations appear in the one-photon and two-photon absorption(OPA and TPA)spectra of FPs,and many other phenomena.Therefore,we select two FPs,LSSmKatel and Citrine,to investigate their spectral properties,and identify the states related to the photophysical and photochemical processes.Meanwhile,we explore the behavior and spectra of aggregation of PBI derivatives.The thesis is organized as follows:Part 1.Characterizing the Structures,Spectra,and Energy Landscapes Involved in the Excited-State Proton Transfer Process of Red Fluorescent Protein LSSmKatel.By applying molecular dynamics(MD)simulations and QM/MM calculations,we have characterized the states and processes involved in the excited-state proton transfer(ESPT)of LSSmKatel.MD simulations identify two stable structures in the electronic ground s-tate of LSSmKatel,one with a protonated chromophore and the other with a deprotonated chromophore,thus leading to two separate low-energy absorption maxima with a large en-ergy spacing,as observed in the calculated and experimentally measured absorption spectra.Proton transfer is induced by electronic excitation.When LSSmKatel is excited,the elec-trons in the chromophore are transferred from the phenol ring to the N-acylimine moiety;the acidity of a phenolic hydroxyl group is thus enhanced.The calculated potential energy curves(PECs)exhibit energetic feasibility in the generation of the fluorescent species in LSSmKatel,and the exact agreement between the calculated and experimentally measured values of the large Stokes shift further provides solid theoretical evidence for the ESPT process taking place in photoexcited LSSmKatel.The molecular environments play a sig-nificant role in the geometries and absorption/emission energies of the chromophores.In addition,TD-?B97X-D/molecular mechanics(MM)provides a better description of the op-tical properties of LSSmKatel than TDB3LYP/MM,although it always overestimates the excitation energies.Part 2.Effect of Protein Environment on One-and Two-Photon Absorption Spec-tra of the Chromophore in the Yellow Fluorescent Protein Citrine.This work is focused on exploring the effect of microenviorment of the protein on OPA and TPA spectra of yellow fluorescent protein Citrine.Starting from the calculation of pure and vibronic OPA and TPA spectra of anionic chromophore(AC)of YFP Citrine and the AC adjunct to the nearby residue Tyr203(AC+Tyr203)in vacuo and solutions,we reveal the intrinsic physical mechanism which results in distinct blue shift of the TPA band relative to the OPA band in region of long wavelength corresponding to the electronic transition from SO to S1.The effects of Herzberg-Teller(HT)vibronic coupling and adjacent residue Tyr203 are responsible for the TPA spectral lineshapes.Beyond the isolated AC and AC+Tyr203 models,then we employ QM/MM method and polarizable embedding density functional theory(PE-DFT)scheme to investigate the explicit effects of protein environment on OPA and TPA spectra of AC+Tyr203.It is found that the residues and water molecules in the vicinity of AC+Tyr203 have different effects on spectra and the main interactions between core system and environment are polarized dipole interactions.To describe the effect of Arg96 correctly,we need a larger model to shield its positively charged guanidinium group.Part 3.Theoretical Investigation on Aggregation of PBI Derivatives Induced by Addition of D-Glucose.To investigate the behavior of aggregation of PBI derivatives due to the addition of D-Glucose in aqueous solution,we perform classical MD simulations,then we calculate the CD spectra of the simplified dimers extracted from the MD snapshots.From MD simula-tions,we found that the spiral directions of PBI-C2-BA and PBI-C4-BA are anticlockwise and clockwise,respectively,indicating that the opposite CD signals.To confirm our guess,we calculated the absorption spectra and CD spectra of a series of dimers which are extract-ed from MD trajectory.The results show that the location of the maximum absorption peak is consistent with experimental value around 500 nm.which originates from the aromatic part of PBI derivatives.The calculated results about CD spectra show that PBI-C2-BA and PBI-C4-BA dimers have the opposite signs and they constitute a pair of enantiomers.By simulating the fold of PBI derivatives and calculating the CD spectra of simplified dimers,we reproduce the experimentally observed phenomena,and conclude that the different spiral manners should be the incentive of CD signals.