VIBRATIONAL CIRCULAR DICHROISM OF SELECTED PEPTIDES, POLYPEPTIDES AND PROTEINS

Dukor, Rina K.

Abstract

Vibrational Circular Dichroism (VCD) has been shown to be a useful method for conformational studies of peptides, polypeptides and proteins in solution. In this thesis, VCD was applied to structural studies of polyproline systems, proline oligomers, model 'random coil' polypeptides and selected proteins.

It was demonstrated that two polyproline structures, polyproline I and II, gave VCD signals differentiable from those of other helices. These two structures have different IR and VCD spectra in solution. Infrared mutarotation studies of PLP I to PLP II in water provided evidence for a two-step transition.

Proline oligomer studies showed that a PLP-II-type helix starts forming at n = 3 and achieves the polymer-like form by n = 5. Based on the chain length dependence of the VCD and UVCD spectra of proline oligomers, it was established that VCD spectra are the result of shorter range interactions than are UVCD spectra.

The VCD of 'random coil' model polypeptides was shown to be identical in shape but smaller in magnitude than that of poly-L-proline II and to have a magnitude similar to that of (Pro)_n, n = 3,4. Based on the spectral evidence, it was concluded that these 'random coils' have a large component of locally conformationally restricted regions. These segments have a conformation similar to the left-handed, 3₁, polyproline II helix, as was previously suggested by Krimm and co-workers. This conclusion is further supported by studies of the effects of salt, temperature and pH on the VCD spectra of L-proline oligomers, poly-L-proline II and poly-L-glutamic acid. After each of these perturbations, a significant local ordering remains in all of the oligomers and polymers studied. Polypeptides such as poly-L-glutamic acid were shown to be more flexible than is poly-L-proline II.

As an application of VCD to the structural study of proteins, the solution conformation of two homologous proteins, alpha-lactalbumin and lysozyme, whose crystal structures are similar, were investigated. Based on the spectral comparison of alpha-lactalbumin and lysozyme, it was concluded that the solution structure of alpha-lactalbumin is not the same as the crystal structure. In particular, the $alpha$-lactalbumin solution structure must have significantly less helix content than does the crystal structure.