Inaug.-diss.--Tübingen. / Vita. "Literaturverzeichniss": p. 28.

Inaug.-diss.--Tübingen. / Vita. "Literaturverzeichniss": p. 28.

Thesis (Ph. D.)--University of Wisconsin--Madison, 1928. / Typescript. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

This work was devoted to the study of (a) the physico-chemical, functional, and structural properties of bean (Phaseolus sp.) protein isolates in relation to their microstructures, and (b) the effects of protein carbohydrate interactions on physico-chemical, functional, and structural properties. The contents of protein, and both total and individual sugars of alkali extracted (amorphous) and citric acid extracted (bipyramidal and spheroidal) proteins from Phaseolus vulgaris (white kidney and navy) and Phaseolus lutanus (baby lima and large lima) beans were determined. The proteins were subjected to differential scanning calorimetry, and measurements of surface tension (air-water interface), surface hydrophobicity, and foam expansion. Structural analysis of the proteins were performed using Fourier transform infrared (FT-IR) spectroscopy. Enzymatic and chemical deglycosylation was performed on a white kidney bean protein isolate. / Glucose and mannose were the major sugars found in the isolates. Bipyramidal and spheroidal microstructures with higher protein contents generally had greater mannose content and lower glucose content. Differences in enthalpy of denaturation $( Delta$H), surface tension decay curves, surface hydrophobicities, and foam expansions were observed with isolates of different microstructures. Corresponding differences in molecular structure were not, however, detected by FT-IR spectroscopy. Using statistical analysis, a relationship between foam expansion and the $ Delta$H, solubility, surface hydrophobicity and surface tension of the isolates was obtained. Preliminary results suggest that the removal of carbohydrate influenced the physico-chemical properties of the protein.

No description available.

The storage reserves in the mature seeds of eleven species of Pinus were investigated. Lipids and proteins 1 sequestered in lipid vesicles and protein bodies respectively 1 were found to be the major storage reserves of pine seeds. All seed tissues of mature Pinus seeds contained protein bodies with one or more protein crystalloids andjor one or more globoid crystals. Energy dispersive x-ray (EDX) analysis of globoid crystals in all species of Pinus that were studied revealed the presence of P 1 K and Mg 1 a fact that is consistent with globoid crystals being phytate-rich. Traces of Ca and Fe were also detected in the globoid crystals of some seed tissues. High levels of Fe and significant levels of P1 K and Mg were detected in small (often .:::; 0. 3 3 J,£m) 1 naturally electron-dense particles that were distributed throughout the tissues of the embryo and female gametophyte. Unlike conventional phytate-rich globoid crystals 1 these Fe-rich particles were not contained in the proteinaceous matrix of typical protein bodies. Instead 1 the particles were contained within membrane-bound structures resembling plastids. Neutron activation analysis and spectrophotometric phosphorus analysis were used to provide a quantitative determination of elements in whole female gametophyte and embryo tissue samples. High levels of P, K, Mg and S and significant levels of Cl, Ca, Mn, Zn and Fe were detected in pine seed tissues. Environment and growth conditions under which the seed developed did not appear to affect the total mineral nutrient levels of mature pine seeds. Seed size and weight varied greatly between the eleven species of Pinus investigated. A major focus of this study was to determine whether seed size was correlated to the distribution of minerals in pine seeds. Previous studies showed that mineral distribution was related to seed size in cucurbits. Phosphorus, Mg and Ca concentrations in globoid crystals and Fe concentrations in electron-dense particles of Pinus seed tissue were found to be negatively correlated with seed size. A comparative study of nutrient reserves in haploid female gametophyte tissue versus diploid embryo tissues was made. Within each species of Pinus, the total mineral nutrient concentrations in whole female gametophyte samples were similar to total mineral nutrient concentrations within whole embryo samples. Protein body structure and the types of mineral nutrients stored in globoid crystals of protein bodies were also similar between female gametophytes and their corresponding embryos. / Thesis / Master of Science (MSc)

The structure of cytochrome c₅₅₅ from the green sulfur bacterium Chlorobium thiosulfatophilum was determined by using a single isomorphous derivative, K₂HgI₄, in combination with its anomalous signal. The initial 2.25 angstrom map was modified by the technique of Fourier inversion. The smoothing function for the electron density map addressed three different features in the map, the solvent density, the protein density and the volume surrounding the heavy atom binding sites known to contain spurious peaks. This structure determination was undertaken for three reasons. First, Chlorobium thiosulfatophilum is a very primitive sulfur metabolizing bacterium and so its cytochrome c₅₅₅ structure is important for its evolutionary implications. Second, the oxidation-reduction potential of cytochrome c₅₅₅ is significantly different from the oxidation-reduction potential of other cytochromes whose structures have been determined. Comparisons with the other structures would provide information concerning the factors that are important in regulating oxidation-reduction potentials. Finally, the three dimensional structure may aid in explaining the pattern of reactivity cytochrome c₅₅₅ displays with mitochondrial cytochrome c oxidase and reductase, which is reversed when compared to other bacterial c-type cytochromes. The resulting structure contains three alpha helices. These features are consistent with other c-type cytochrome molecules previously determined. Two regions of the map appear to be disordered and are difficult to interpret. Possible causes of this observation are discussed and related to the significance of the structure.

Fluid resuscitation with crystalloid and colloid solutions is a common treatment in perioperative medicine. However, a variety of unbalanced or balanced solutions is used in clinical practice and there is still a vivid debate going on regarding the selection of optimal fluid with minimal negative effect on coagulation. The goal of the dissertation was to investigate the adverse effect of balanced crystalloids and colloids on whole blood coagulation measured by method of rotational thromboelastometry. In the first phase of the work we had assessed the adverse effect of balanced crystalloid, hydroxyethyl starch and gelatin after dilution of blood with the solution in vitro. Parametrs of EXTEM and FIBTEM tests were evaluated by using rotational thromboelastometry. In the second phase of the work we evaluated the negative effect of infusion solution after dilution in vivo. We had obtained blood samples from 30 patients during knee arthroscopy before and after administration of 500 ml of crystalloid or hydroxyethyl starch or gelatin. Parametrs of EXTEM and FIBTEM tests were evaluated by using rotational thromboelastometry.In compliance with the results of the dissertation, hydroxyethyl starch has the most obvious negative effect on clot formation followed by gelatin and finally by crystalloids. Based on...

X-ray crystallography is a popular method for resolving protein structures. Protein crystals need to be used for X-ray crystallography, but most naturally occurring proteins do not readily crystallize. The Hunt lab performed computational analyses showing that arginine is the most overrepresented amino acid in crystal-packing interfaces in the Protein Data Bank. Given the similar physicochemical characteristics of arginine and lysine, we hypothesized that multiple lysine-to-arginine (KR) substitutions should improve crystallization. To test this hypothesis, we developed software that ranks lysine sites in a target protein based on the redundancy-corrected KR substitution frequency in homologs. We demonstrate that three unrelated single-domain proteins can tolerate 5-11 KR substitutions with at most minor destabilization and that these substitutions consistently enhance crystallization propensity. This approach rapidly produced a 1.9 Å crystal structure of a human protein domain refractory to crystallization with its native sequence. Structures from bulk-KR-substituted domains show the engineered arginine residues frequently make high-quality hydrogen-bonds across crystal-packing interfaces. We thus demonstrate that bulk KR substitution represents a rational and efficient method for probabilistic engineering of protein surface properties to improve protein crystallization. This stands in direct contrast to earlier work and dogmas that posited that surface entropy reduction was the clear path forward to crystallzing proteins. Arginine is a high-entropy sidechain, yet it helps drive protein crystallization. To understand which structure and dynamical features of arginine give rise to crystal packing propensity, we performed 60 Molecular Dynamics (MD) simulations to measure the sidechain order parameter of arginine and compare it against crystal packing propensity. This work found that surface-exposed arginines with low order parameters are most likely to participate in crystal packing interactions. This is evidence against earlier thinking that high entropy surface sidechains oppose crystallization. Entropic barriers to protein crystallization can be enthalpically overcome.

This microcirculatory study compared the effects on oxygen transport of two hemodilution fluids: HBOC-201 (Biopure Corp., Cambridge, MA) a Hemoglobin-Based Oxygen Carrier (HBOC), and 5.9% Human Serum Albumin (HSA) an iso-oncotic non-oxygen carrying colloid solution. Measurements using intravital microscopy were made on the spinotrapezius muscle of male, Sprague-Dawley rats. Interstitial PO2 was measured using phosphorescence quenching microscopy, and recorded before and after isovolemic hemodilutions (HD) at hematocrits of 40% (baseline), 30% (moderate HD) and 15% (severe HD). Oxygen consumption (VO2) of the spinotrapezius muscle was derived from PO2 recordings following the rapid inflation of a plastic bag placed around the objective. When the bag was inflated, blood flow in the muscle was arrested and PO2 rapidly fell over several seconds; the rate of decline of PO2 was proportional to VO2. For moderate HD (Hct ~ 30%) with HBOC-201, interstitial PO2 did not change from baseline conditions (Hct ~ 40%), while HD with HSA showed a decrease. For severe HD (Hct ~ 15%) both PO2 and VO2 were significantly lower for the HSA group than for the HBOC-201 group. These findings indicate that HBOC-201 maintains both a higher PO2 and VO2 during hemodiluted states compared with a non-oxygen carrying colloid solution (HSA). Furthermore, 5.9% HSA does not affect the mean arterial pressure (MAP) and vessel diameters, whereas HBOC-201 causes vasoconstriction, and consequently an increase in MAP. However, the vasoconstriction is not uniform among different branches of the arteriolar network, and most of the changes occur in the larger vessels, i.e., feed and arcade arterioles, while minimal in smaller vessels, i.e., transverse arterioles. In addition, findings show that MAP and vessel diameters return to baseline within 1-3 hours, implying that vasoconstriction and hypertension caused by HBOC-201 are acute responses.