Lednev Igor K, Ermolenkov Vladimir V, He Wei, Xu Ming
Department of Chemistry, University at Albany, SUNY, Albany, NY 12222, USA.
Anal Bioanal Chem. 2005 Jan;381(2):431-7. doi: 10.1007/s00216-004-2991-5. Epub 2004 Dec 30.
A new deep-UV Raman spectrometer utilizing a laser source tunable between 193 and 205 nm has been designed, built, and characterized. Only selected wavelengths from this range have previously been accessible, by Raman shifting of the second, third, and fourth harmonics of the Nd:YAG fundamental in hydrogen. The apparatus was demonstrated to be a useful tool for characterizing hen egg white lysozyme structural rearrangements at various stages of fibril formation. High-quality deep-UV resonance Raman spectra were obtained for both a protein solution and a highly-scattering gelatinous phase formed by fibrillogenic species. In addition to amide bands, strong contribution of nu(12) and ring-C phenylalanine vibrational modes was observed at excitation wavelengths below 200 nm. Remarkably, the Raman cross-section of these modes revealed dramatic change of lysozyme in response to heat denaturation and fibril formation. These results indicate that phenylalanine could serve as a new deep-UV Raman probe of protein structure.
一种新型的深紫外拉曼光谱仪已被设计、制造并进行了特性表征，该光谱仪利用了一个波长可在193至205纳米之间调谐的激光源。此前，通过在氢气中对Nd:YAG基频的二次、三次和四次谐波进行拉曼频移，只能获得该范围内的特定波长。该仪器被证明是一种用于表征蛋清溶菌酶在纤维形成不同阶段结构重排的有用工具。对于蛋白质溶液和由纤维形成物种形成的高散射凝胶相，都获得了高质量的深紫外共振拉曼光谱。除了酰胺带外，在低于200纳米的激发波长下，还观察到了ν(12)和环-C苯丙氨酸振动模式的强烈贡献。值得注意的是，这些模式的拉曼截面显示出溶菌酶在热变性和纤维形成时的显著变化。这些结果表明，苯丙氨酸可以作为一种新的蛋白质结构深紫外拉曼探针。
