Backbone ‘Stretched;’ Backbone ‘Curled’: Conformational Similarity for Trp Dipeptides Parallels Fluorescence Emission and UVRR W10 Shifts

Abstract

Tryptophan is a spectroscopic marker of protein environment because the fluorescence emission maximum of its residue, indole, is sensitive to both solvent and the immediate protein environment. The ease of measuring indole fluorescence accounts for the popularity of characterizing proteins via this spectroscopic technique. However, molecular level detail that accounts for emission shifts is lacking. Here we address the problem of emission shifts caused by amino acid conformational changes in aqueous solution by examining a variety of tryptophan dipeptides in the zwitterionic and anionic states. In addition to fluorescence emission, we employ UV resonance Raman (UVRR) spectroscopy, molecular dynamics simulation, and ab initio calculations. In our previous study of TrpGly and GlyTrp dipeptide species, a correlation between fluorescence emission shifts and UVRR W10 band shifts and W7 band intensity ratios were discovered. Nanosecond molecular dynamics simulation coupled with energy minimization showed that the dipeptide species adopt two basic conformations: one with a ‘stretched’ backbone or one with a ‘curled’ backbone. For TrpGly zwitterions in the stretched conformation, the terminal amine cation is 2.44 A from the pyrrole ring C3. The Stark effect predicts that a positive charge near the pyrrole ring blue shifts the emission maximum, which is observed. Remarkably, rotamers do not show a correlation with the dihedral angle, chi 2. Rather, the dihedral angle, chi 1, takes on three discrete values that account for the ‘stretched’ and ‘curled’ backbone conformations. We have continued in this manner to study Trp dipeptides with Glu, Lys, Tyr, His and Trp, showing that both the pattern of backbone interaction with the indole and the spectroscopic pattern of changes with dipeptide species persists in spite of residue variation in charge, aromaticity and size.