Laser Raman spectroscopic studies of ocular lens and its isolated protein fractions

The water soluble proteins of the bovine lens were separated on a column of Sephadex G 200 into 5 fractions designated as α, β₁, β₂₇, B₃, and γ crystallin. Laser Raman scattering studies on these isolated proteins (both in the lyophilized state and in solution), and insoluble albuminoid, reveal that they contain predominantly antiparallel pleated sheet structure in the main chains, and that sulfhydryl groups are highly localized in γ crystallin. This light scattering technique was also applied to probe the homogeneity of protein strucutre in the intact lens. The analysis of the scattered light selectively collected from various parts of the lens indicates that these proteins also exist in an antiparallel β structure throughout the entire lens. However, the central (nucleus) and outer (cortex) portions have somewhat different amino acid composition. Based on the relative intensities of the lines at 624 (phenylalanine) and 644 cm-¹ (tyrosine), it is concluded that the nuclear part has the highest concentration of γ crystallin, and that the content of α crystallin increases significantly from the nucleus to the cortex. By examining the Raman spectra in the 2582 cm-¹ and the amide I and III regions, the authors have demonstrated that the sulfhydryl groups and the β conformation of the lens proteins are unaffected in the conversion of transparent to totally opaque lens by heat denaturation at 100°. This means that the opacification of a lens does not necessarily involve the oxidation of sulfhydryl groups or conformation changes.