Patrik R. Callis

We are currently studying the internal structure and dynamics of proteins through a combination of molecular modeling, quantum mechanics, and ultraviolet spectroscopy. We are primarily exploiting the amino acid tryptophan, whose spectra are sensitive to the internal electric field caused by the surrounding protein scaffolding and solvent. We have already a good quantum mechanical understanding of the tryptophan excited states, one that strongly suggests that the wavelength of the fluorescence emitted from the excited state after excitation with UV light depends almost entirely on the magnitude and direction of the electric field caused by charges and dipoles of the protein and solvent. Current focus is on the more challenging problem of understanding the basis of the 30-fold variation in fluorescence quantum yield found for tryptophans in different protein environments. By combining quantum mechanics and molecular dynamics simulations to generate parameters for electron transfer rate theory, we have found a promising correlation between quantum yield and the computed energy gap between the emitting state and a higher lying charge transfer excited state. The significance of this work lies in the fact that hundreds of experiments are published every year exploiting the changes in tryptophan fluorescence in proteins for a variety of studies, including protein folding pathways and dynamics, enzyme action, and molecular recognition. It is essential that this probe be understood at a fundamental level, because the ultimate detailed pictures of proteins cannot be established from conventional structure methods alone. In addition, if the electrostatic model we have proposed proves correct, tryptophan fluorescence wavelengths will provide excellent tests for the important task of accurately predicting electrostatic fields and potentials in proteins.

Quantitative prediction of fluorescence quantum yields in proteins: Tryptophan

Tryptophan fluorescence intensity is widely used to monitor almost any imaginable change in protein structure. Tryptophan fluorescence intensity is widely used to monitor almost any imaginable change in protein structure. Using hybrid quantum mechanics-molecular mechanics (QM-MM) simulations we have recently shown that the full 30-fold range of Trp fluorescence quantum yields (and lifetimes) observed in proteins is due primarily to different rates of electron transfer from the excited indole ring to one of two nearest backbone amides. This heretofore puzzling dependence on protein environment arises mainly from the average local electric potential difference between the Trp ring and acceptor amide and from the amplitude of potential difference fluctuation caused by protein and solvent motions.

  1. We continue to pursue stronger documentation of the precise nature of the charge transfer state on the amide through higher level quantum calculations.
  2. We continue to apply our method to test and predict Trp fluorescence in a growing array of proteins.
  3. We are refining similar predictions for quenching caused by other protein residues, including protonated histidine, disulfide, amide side chains, and cysteine.
  4. We are studying the nature of the electron transfer quantum mechanical matrix element, which is a major factor in the quenching process.

Selected Publications:

Callis PR, Liu T:
Quantitative predictions of fluorescence quantum yields for tryptophan in proteins
J. Phys. Chem. B 108 4248-4259 (2004)

Xu J, Toptygin D, Graver KJ, Albertini RA, Savtchenko RS, Meadow ND, Roseman S, Callis PR, Brand L, Knutson JR :
Ultrafast Fluorescence Dynamics of Tryptophan in theProteins Monellin and IIAGlc
J. Am. Chem. Soc. 128 ASAP (2006)

Callis PR, Vivian JT:
Understanding the variable fluorescence quantum yield of tryptophan in proteins using QM-MM simulations. Quenching by charge transfer to the peptide backbone
Chem. Phys. Letters 369 409-414 (2003)+A49

Keywords:

Biophysical, Protein Chemistry

Quantitative prediction of fluorescence quantum yields in proteins: Flavins and dyes

Oxidized flavin cofactors exhibit an even wider range of fluorescence intensities and lifetimes than tryptophan. The source of this quenching is better understood: flavin quenching is almost always by electron transfer from nearby unexcited tryptophan and tyrosine to the excited state of the flavin. The same appears to be the case for many of the fluorescent dyes that are attached to proteins for various imaging and diagnostic purposes. We are modeling the efficiency of this process with the same tools we use for modeling Trp fluorescence.

Selected Publications:

Callis PR, Liu T :
Short Range Photoinduced Electron Transfer in Proteins: QM-MM Simulations of Tryptophan and Flavin Fluorescence Quenching in Proteins
Chemi Phys. (2006) in press+A60+A25

Keywords:

Biophysical, Protein Chemistry

Quantitative prediction of fluorescence spectra and spectral relaxation of tryptophan in proteins.

The tryptophan fluorescence spectrum in proteins depends sensitively on the local environment covering the range 308-355 nm. We have previously shown that the wavelength can be predicted from QM-MM simulations. One must sum over all charged atoms of the protein and solvent to find the electric potential different from the 5- to the 6-membered ring of the Trp. Recently, there is considerable interesting in how rapidly the solvent and protein environment responds to the suddent large dipole increase caused by excitation of Trp. We are modelling the dynamics of this shift with our QM-MM simulations to see why controversial, long relaxation times appear in some experiments.

Selected Publications:

Xu J, Toptygin D, Graver KJ, Albertini RA, Savtchenko RS, Meadow ND, Roseman S, Callis PR, Brand L, Knutson JR:
Ultrafast Fluorescence Dynamics of Tryptophan in theProteins Monellin and IIAGlc
J. Am. Chem. Soc. 128 ASAP (2006)

Vivian JT, Callis PR:
Mechanisms of Tryptophan Fluorescence Shifts in Proteins
Biophys. J. 80 2093-2109 (2001)

 

Keywords:

Biophysical