Indexed on: 02 May '07Published on: 02 May '07Published in: Analyst
Rich and complex Raman scattering and Raman optical activity (ROA) spectra have been measured monitoring the pH induced alpha-helix-to-disordered conformational transition in poly(L-glutamic acid). Two-dimensional (2D) correlation techniques have been applied to facilitate a comprehensive analysis of these two complementary spectral sets. Synchronous contour plots have identified band assignments of alpha-helical and disordered conformations, and have revealed bands characteristic of changes in the protonation state of the polypeptide. Asynchronous plots, on the other hand, have probed the relative sequential orders of intensity changes indicating a decrease in intensity of alpha-helical bands in the backbone skeletal stretch region, followed by a subsequent decrease in intensity in the extended amide III and amide I regions, underlying the appearance of disordered structure, including poly(L-proline) II (PPII) helix. The application of a 2D correlation 'moving' window has also disclosed two distinct phases during helix unfolding in the alpha-helix-to-disordered transition, occurring at approximately pH 4.9 and approximately pH 5.2, possibly a result of the difference in helical stability between the end and central regions of the alpha-helix. This paper demonstrates the potential value of combining 2D Raman, 2D ROA and moving window correlation techniques for the detailed investigation of complex and subtle changes of secondary structure during the unfolding mechanisms of polypeptides and proteins.