Difference FTIR Spectra Research Articles

Spectroscopic Studies of Oxidative Degradation of Base Oils

NMR and FTIR spectroscopies have been used for obtaining component-level information on the thermo-oxidation of some base oils. Base oils were first separated by column chromatography into their saturates, aromatics, and polars. These fractions as well as the neat base oil were oxidized by a modified IP 306 benchtop (simulated) method. A 1H and 13C NMR-derived structural model was utilized to monitor the structural changes and rearrangements occurring in the carbon skeleton of the hydrocarbons during oxidation of base oils/fractions. FTIR was used to measure variations in the concentration profiles in the carbonyl absorption region (1649−1815 cm-1). Difference FTIR spectra and deconvolution techniques were employed to detect new compounds formed as well as minor concentration changes in oil components. NMR and FTIR results were utilized to study the role of different hydrocarbon types in the oxidation of base oil and to establish the reaction mechanism responsible for the degradation of base oils.

Water and Peptide Backbone Structure in the Active Center of Bovine Rhodopsin

Difference FTIR spectra in the conversion of rhodopsin or isorhodopsin to bathorhodopsin were recorded for recombinant wild-type and E113Q bovine rhodopsins. Differences in various vibrational modes between E113Q and the wild-type proteins whose Schiff bases interact with chloride and Glu113, respectively, were analyzed. Water molecules in rhodopsin that change upon formation of bathorhodopsin are detected by a change in frequency of the O-H stretching vibration from 3538 to 3525 cm(-1). This change in the wild-type protein is absent in E113Q. One or a few water molecules are therefore suggested to be located in the proximity of Glu113, the counterion of the Schiff base. Another water vibration at 3564 cm(-1), which is shifted to 3542 cm(-1) in bathorhodopsin in the wild type, persists in E113Q but with approximately 5-cm(-1) shift toward higher frequency. This is due to water molecules that may be located at a site somewhat more remote from Glu113. Structural changes of some peptide carbonyls and amides are also absent in E113Q. On the other hand, the E113Q protein shows shifts of the N-H+ stretching vibrational band, that is probably due to the protonated Schiff base, upon conversion of rhodopsin to bathorhodopsin. No corresponding changes were observed in the wild type. We propose a model in which a water molecule interacts with Glu113, the protonated Schiff base, and peptide carbonyls, and amides. These residues undergo structural changes upon formation of bathorhodopsin.

The structure and interactions of Ca(2+)-ATPase.

Electron crystallographic studies on membrane crystals of Ca(2+)-ATPase reveal different patterns of ATPase-ATPase interactions depending on enzyme conformation. Physiologically relevant changes in Ca2+ concentration and membrane potential affect these interactions. Ca2+ induced difference FTIR spectra of Ca(2+)-ATPase triggered by photolysis of caged Ca2+ are consistent with changes in secondary structure and carboxylate groups upon Ca2+ binding; the changes are reversed during ATP hydrolysis suggesting that a phosphorylated enzyme form of low Ca2+ affinity is the dominant intermediate during Ca2+ transport. A two-channel model of Ca2+ translocation is proposed involving the membrane-spanning helices M2-M5 and M4, M5, M6 and M8 respectively, with separate but interacting Ca2+ binding sites.

The effect of dicyclohexycarbodiimide and cyclopiazonic acid on the difference FTIR spectra of sarcoplasmic reticulum induced by photolysis of caged-ATP and caged-Ca2+

The photochemical release of Ca2+ from caged-Ca2+ in the absence of ATP, and the release of ATP from caged-ATP in the presence of Ca2+ induce characteristic difference FTIR spectra on rabbit sarcoplasmic reticulum that are related to the formation of Ca2E1 and E∼P intermediates of the Ca2+-ATPase, respectively. Dicyclohexylcarbodiimide (10 nmol/mg protein) abolished both the Ca2+-and ATP-induced difference FTIR spectra parallel with inhibition of ATPase activity. Cyclopiazonic acid (50 nmol/mg protein) inhibited the Ca2+-induced difference spectrum measured in the absence of ATP, but had no significant effect on the ATP-induced difference spectrum measured in the presence of 1 mM Ca2+. The dog kidney Na+,K+-ATPase did not give significant difference spectrum after photolysis of caged-ATP in Ca2+-free media containing 90 mM Na+ and 10 mM K+, with or without ouabain. We propose that both the Ca2+ and the ATP-induced difference FTIR spectra of the Ca2+-ATPase reflect the occupancy of the high-affinity Ca2+ transport site of the enzyme.