Protein simulations: The absorption spectrum of barnase point mutants

Abstract

The near-UV absorption spectra of barnase double-point mutants are calculated using a combination of molecular dynamics and ab initio techniques. The atoms of the fluorescent probes are placed in a cloud of point charges, generated by molecular dynamics simulations. Ab initio calculations (CASPT2) are performed on these systems. Three molecular dynamics packages are compared-Amber5.0, CHARMMc27b1, and GROMOS96-using indole as the fluorescent probe. It was found that calculated absorption spectra reproduce experimental values very well, provided detailed charge cloud descriptions are included. These calculations further sustain the hypothesis that different tryptophan rotamers can be present in proteins. Molecular dynamics calculations of the double-point mutants also point to the structural effect of counter ions.

Modeling the Absorption Spectrum of Tryptophan in Proteins

“…The calculated gas-phase (isolated) indole transition energies are nearly identical for each of the three mutants. Upon addition of just the extended environment represented by point charges (R ) 0.0 Å), the 1 18 whose best results (using an AMBER force field and charges with snapshot geometries) are 4.32 and 4.73 eV for transitions to 1 L b and 1 L a , respectively. These correspond to red shifts of 0.04 and 0.07 eV with respect to vertical transition energies calculated in the gas phase.…”

“…18 Wild-type barnase has three Trp residues. The three Trp double point mutants will each leave a different single Trp in the protein, thereby allowing the spectrum of the remaining Trp to be studied.…”

“…In previous work, this led to only the chromophore itself being included explicitly within the excited-state quantum-chemical calculation, with the extended environment modeled using point charges. 18 This close proximity of a large number of point charges in the CASSCF/CASPT2 calculations may be partially responsible for the large variations observed between the different force fields. In calculations of electronic spectra in solution there are many instances of solvatochromic shifts that are not described by electrostatics only.…”

Porcine pancreatic alpha amylase and its isoforms—Effect of deglycosylation by peptide-N-glycosidase F

“…The absorbance is slightly more in deglycosylated samples when compared to the glycosylated enzyme in buffer. These absorption spectra are also sensitive to the local protein environment, but, however, less pronounced compared to the fluorescence spectra [25].…”

“…absorption spectra of glycosylated and deglycosylated derivatives of PPA and its isoforms in sodium phosphate buffer (pH 6.9, 0.02 M) were recorded using Shimadzu UV-160 spectrophotometer in a wavelength range of 200-400 nm [25].…”