DESIGN AND PERFORMANCE OF AN ULTRAVIOLET RESONANCE RAMAN SPECTROMETERFOR PROTEINS AND NUCLEIC-ACIDS

We describe an ultraviolet resonance Raman (UVRR) spectrometer appropr iate for structural studies of biological macromolecules and their ass emblies. instrument design includes the following features: a continuo us wave, intracavity doubled, ultraviolet laser source for excitation of the Raman spectrum; a rotating cell (or jet source) for presentatio n of the sample to the laser beam; a Cassegrain optic with f/1.0 apert ure for collection of the Raman scattering; a quartz prism dispersing element for rejection of stray light and Rayleigh scattering; a 0.75-m single grating monochromator for dispersion of the Raman scattering; and a liquid-nitrogen-cooled, charge-coupled device for detection of t he Raman photons. The performance of this instrument, assessed on the basis of the observed signal-to-noise ratios, the apparent resolution of closely spaced spectral bands, and the wide spectrometer bandpass o f 2200 cm(-1), is believed superior to previously described UVRR spect rometers of similar design. Performance characteristics of the instrum ent are demonstrated in UVRR spectra obtained from standard solvents, p-ethylphenol, which serves as a model for the tyrosine side chain, th e DNA nucleotide deoxyguanosine-5'-monophosphate, and the human tumor necrosis factor binding protein, which is considered representative of soluble globular proteins.