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[en] Research is reported on magnetic resonance spectroscopy of biological molecules, development of clinical applications of stable isotopes, circadian cybernetics, and X-ray crystallography of immunoglobulins. Biological processes occur in fluid media, and ultimately our knowledge of their mechanisms requires detailed information for chemical and molecular structural properties in biological fluids. Magnetic resonance spectroscopy has unique advantages over other approaches in this area that are being exploited in studies currently underway in the group. The program continues to develop along three interrelated lines, measurement and analysis of high resolution spectra for biological molecules (especially nucleic acid constituents and drugs), synthesis of selectively labeled nucleic acid fragments essential for complete spectral assignments, and computation of conformational properties from NMR parameters. This coordinated approach enabled the first complete conformation analysis for a dinucleoside monophosphate, ApA, in aqueous solution. It was found that the conformation is actually a time-average of right helical, loop, and extended conformations, the interchange being extremely rapid on an NMR time scale. Spectral analyses were also completed for all possible ribonucleotide dimers, the assignments again relying heavily on synthesis of appropriate deuterated counterparts. Studies of conformational flexibility in nucleic acid fragments showed that changes in hydrogen ion concentration and temperature produce correlated conformational changes specific for each nucleotidyl unit. Studies were also initiated in three new projects dealing with the effect of hapten binding on antibody structure, counter ion influence on nucleic acid free radicals, and membrane differences between normal and sickled erythrocytes

[en] This section contains summaries of research on the detection and characterization of damage in molecular, cellular, and physiological systems. Projects under investigation in this section include: chemical synthesis of nucleic acid derivatives; structural and conformational properties of biological molecules in solution; crystallographic and chemical studies of immunoglobulin structure; instrument design and development for x-ray and neutron scattering studies of biological molecules; and chromobiology and circadian regulation

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[en] The objectives of this project are to develop instrumentation for small angle x-ray and neutron scattering, and to utilize small angle techniques for study of the structures of the intracellular molecules interacting with the secondary messengers involved in cellular regulation. A unique self-scanning photodiode array has been developed as a linear position sensitive detector for studies of biological structures. A time-of-flight (TOF) small angle neutron instrument was developed and successfully tested at the prototype pulsed neutron facility, ZING-P'. Considerable hardware and software developments were necessary to successfully demonstrate the prototype small angle neutron scattering instrument. A dedicated data acquisition system based on a microprocessor was developed and tested within the short period of approximately 6 months and was interfaced to a biological sample changer and environmental controller. The resolution of the tapered collimation system proved to be adequate

[en] The first detailed study has been made of the 220-MHz NMR spectra of α-adenosine 3',5'-cyclic monophosphate (I,α-cAMP), α-uridine 3',5'-cyclic monophosphate (II, α-cUMP), α-cytidine 3',5'-cyclic monophosphate (III, α-cCMP), α-deoxyadenosine 3',5'-cyclic monophosphate (IV, α-cdAMP), α-5,6-dihydrouridine 3',5'-cyclic monophosphate (V, α-cDHUMP), and β-5,6-dihydrouridine 3',5'-cyclic monophosphate (VI, β-cDHUMP) in D₂O solution. Analyses of the spectra of I-III were aided by the use of europium chloride as a shift reagent. A conformational analysis showed the sugar moieties of I-III to exhibit a conformation in the range ₂T³ to ³T₂ with an unusually high distortion from planarity, in contrast to the β anomers which prefer ³E to ₄T³ and the acyclic mononucleotides which show a ²E reversible ³E equilibrium. This change in the preferred conformation is attributed to a repulsive interaction between the 2'-hydroxyl and the base. Removal of the 2'-hydroxyl group eliminates this interaction and causes a relaxation to a less strained system. This is clearly demonstrated in the sugar ring conformation of IV which exhibits a ³E to ₄E pucker and a puckering amplitude that is less than in the ribo series. Hydrogenation of the pyrimidine ring of II and β-cUMP gave the 5,6-dihydro products V and VI. VI exhibits preference for a ³E to ₄T³ ribose ring conformation and in the case of V the 2'-hydroxyl-base interaction is markedly reduced owing to the increased flexibility of the aglycon. This results in a relaxation of the sugar ring conformation from the ₂T³ to ³T₂ in I-III back toward the ³E to ₄T³ conformation found in the β anomers. Saturation of the base is not as effective as removal of the 2'-hydroxyl group in relieving the strain in these rigid systems. The phosphate ring is found to be in a flattened chair form in all cases

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[en] ¹³C Nuclear magnetic resonance chemical shifts, ¹J/sub c-c/ scalar coupling constants, spin-lattice relaxation times, and nuclear Overhauser effects were determined for taurine-[1, 2 ¹³C] and a taurine-[1 ¹³C] and taurine-[2 ¹³C] mixture in the presence and absence of calcium. Comparison of taurine titration shifts to values for related compounds reveals some unusual electronic properties of the taurine molecule. Stability constants of 1:1 calcium complexes with taurine zwitterions and anions, as well as their ¹³C chemical shifts, were obtained by least squares analysis of titration curves measured in the presence of calcium. The stability constants of calcium-taurine complexes were significantly lower than previous values and led to estimates that only approximately one percent of intracellular calcium of mammalian myocardial cells would exist in a taurine complex

[en] A linear position sensitive small-angle x-ray detector has been developed which utilizes a directly exposed self-scanning photodiode array cooled to liquid nitrogen temperature to reduce fixed pattern noise. Direct exposure of the array improves the overall sensitivity to 8 keV photons by an order of magnitude over photoconversion techniques. With this improvement in sensitivity, the photodiode array has been effectively used to detect x rays from weakly scattering biological molecules in solution. The slit-like geometry of the photodector elements ideally matches that of a Kratky collimator employed with a low-flux x-ray source to enhance greatly the data collection rate. The detective quantum efficiency and stability of the detector have been determined while taking small-angle scattering measurements of proteins and the results are quantitatively applied to characterize the performance of the instrument