1H NMR spectroscopic studies on bovine lens crystallins

The research in this thesis involved the separation and isolation of crystallin proteins from bovine lenses and the analysis of the protein isolates by 1 H two dimensional NMR spectroscopy. Sequential assignments were made for extended and flexible terminal regions of βB2-, γs- and γB-crystallin, which assume random coil conformations. NMR spectral analysis showed that different conformations of the terminal extensions and flexible regions within the domain regions exist between the p subunits. This is consistent with separate roles for each subunit in aggregate formation. It is shown that the C-terminal extension of the βB2 subunit is involved in total β aggregation at in vivo concentrations. Evidence is also presented to show that the γs-crystallin has conformational attributes intermediate to those of the β- and γ-crystallins. NMR spectra acquired for a range of crystallin homogenates give supporting evidence for supra-molecular interactions between the crystallins with specific interactions deduced for the assigned terminal extensions. Based on these results a model is proposed of how the crystallins maintain short order interactions consistent with lens transparency.