Graduation date: 1969

The results of a preliminary experiment suggested that a complexed form of cobalt was more efficacious in promoting growth of a marine phytoplankton than ionic cobalt. The phytoplankton used in this experiment was Coccolithus huxleyi, a vitamin B₁₂ producer, and the cobalt complex was cobalt (II)- ethylenediaminetetracetic acid [Co(II)- EDTA]. A review of the biochemistry of vitamin B₁₂ indicates that a B₁₂ producer might prefer, if not require, Co(III) instead of Co(II). Since some of the Co(II)-EDTA in the preliminary experiment might have become oxidized to Co(III)-EDTA, the observed stimulation of growth could have been due to Co(III)-EDTA. Two experiments were performed to determine if Co(III)-EDTA is more efficacious in stimulating growth than Co(II)-EDTA. Coccolithus huxleyi, grown in batch cultures with constant illumination, was used in both experiments. One experiment had cobalt concentrations of 10 and 1 μg/l; the other had concentrations of 1 and 0.1 μg/1. In both experiments there were no observed differences in specific growth rates between treatments of Co(III) as the EDTA complex with 10⁻⁶ M additional EDTA and Co(II) with 10⁻⁶ M EDTA. Both of these treatments resulted in a specific growth rate larger than controls without added EDTA or cobalt. It is not possible to measure the amount of Co(II)-EDTA that is oxidized to Co(III)-EDTA at the concentrations used in these experiments. Consequently these results cannot be used as a basis for rejecting the hypothesis that Co(III) is the required form of cobalt. Since Co(III)-EDTA without the additional 10⁻⁶ M EDTA was apparently able to stimulate growth in relation to the controls, it is assumed that C. huxleyi is capable of utilizing this form of cobalt. / Graduation date: 1975

Graduation date: 1991

Specimens of Citharichthys stigmaeus, the speckled sanddab, were taken from Yaquina Bay, Newport, Oregon and exposed for eight days to dissolved measured concentrations ranging from 2.9 μg/l to 190 μg/1 of the polychlorinated biphenyl, Aroclor® 1260. Oxygen consumption was measured to determine any change from routine levels of respiration. This research produced the following conclusions: 1) a relationship exists between oxygen consumption and measured concentrations of Aroclor® 1260 in the water as demonstrated by high negative partial correlations and subsequent significant water concentration terms in the regression equations for two of the four experiments: also, a relationship exists between oxygen consumption and tissue concentration of Aroclor® 1260 as demonstrated by high negative partial correlations and subsequent significant tissue concentration terms in the regression equations for three out of four experiments, (because of the inherent variability in oxygen consumption measurements and difficulties of predicting PCB concentrations in both water and tissue, the evidence is difficult to interprete); 2) acetone, used as a solvent for the toxicant, may change oxygen consumption levels and interfere with determining PCB effects; 3) after an eight day dosing period the concentration of Aroclor® 1260 in whole body tissue can be described by the equation: Y = 4.48(1 - e [superscript -.0365x]) where Y is the concentration of Aroclor® 1260 in the tissue and x is the measured Aroclor® concentration in the water; 4) after eight days a constant level of Aroclor® 1260 is present in the tissue if the animal is exposed to water levels of Aroclor® 1260 above approximately 25 μg/1. / Graduation date: 1976

A detailed study of iron concentrations measured by the reducible, reactive ferrous, and extractable methods was conducted in Auke Bay, Alaska during June 1973 and in Hood Canal, Washington during October 1973 and April 1974. The concentrations by each method were generally highest at the bottom and decreased towards the surface with occasional pockets of high iron concentration water at the surface. In Hood Canal in October the maximum reactive ferrous iron concentration was observed at the bottom of the pycnocline. Large changes in the iron concentration were observed between successive surveys. The reducible iron method (range <0.8 to 187 μg Fe/L) measures colloidal ferric hydroxide plus easily dissolvable and reducible organically and inorganically bound iron. The reactive ferrous iron method (range <0.04 to 4.2 μg Fe/L) measures loosely bound organic and inorganic ferrous iron. The extractable iron method, as finally used, (range <4 to 170 μg Fe/L) measures that iron measured by reducible iron method plus up to twice the tightly bound organic iron. We have defined "available" iron (with quotes) as that iron which appears to be taken up by phytoplankton as determined by the correlation between the in situ iron concentration and the chlorophyll a concentration. Chlorophyll a concentration maxima generally occurred at the depth of the iron concentration minima, strongly supporting the idea that each method measures "available" iron. However, significant correlation coefficients for the regression of iron concentration versus chlorophyll a only support the "availability" of reactive ferrous iron and that phytoplankton prefer ferrous iron to ferric iron. This was one of the first extensive uses of Lewin and Chen's (1973) ferrous iron method and values observed were generally 2 to 100 times less than they reported. Changes occurring during water storage appear to be a function of p0₂. Major changes took place during the first five hours of storage. Interstitial water was greatly enriched in iron compared with the overlying water. The ferrous iron concentration was particularly high due to the low Eh of the sediments. Although the net flow of "available" iron is out of the sediment, the flux is small (approximately 3 x 10⁻⁵ μg Fe/m²sec). Most of the iron that appears to be coming from the sediment is probably being produced by bacterial decomposition as in the case of subsurface sources of iron associated with ammonia or urea maxima. Fresh water is a significant source of extractable and reactive ferrous iron into the well-mixed surface layer. Fresh water is also a source of organically bound iron. Zooplankton appear to produce "available" iron by eating or excretion. Light appears to cause the production of ferrous iron through an undetermined mechanism. / Graduation date: 1976

Investigation was made into the influence of changing light intensity on the organic and elemental composition of two species of marine diatoms, Ditylum brightwellii and Skeletonema costatum. Analysis was made of the relative changes in soluble and insoluble carbohydrate; polar, non-polar, and total lipid; amino acids; proteins; and total carbon, hydrogen, nitrogen, and oxygen. Photosynthetic quotients were calculated from both organic and elemental composition. The response of soluble and insoluble carbohydrate was directly related to light intensity, although differences were noted between the two fractions. In general, amino acids and proteins showed little or no response to light intensity. Polar (pigment) lipids usually varied inversely with light intensity, whereas non-polar lipids varied directly with intensity. Total lipids also varied inversely with intensity although, considering the total amount of lipid present, the changes were slight. Some differences in relative organic composition were noted between the two species. On the basis of the data, average organic composition of each species was computed and compared with similar data in the literature. The effect of light intensity on the elemental composition of the two species yielded some differences, which were more readily apparent in the photosynthetic quotients calculated from elemental data. The photosynthetic quotient varied inversely with light intensity. Conclusions are summarized and suggestions for future research included. / Graduation date: 1970

Nitrate reductase is known to be the enzyme regulating the reduction of nitrate to nitrite in plants. This reduction is the first and rate-limiting step in the transformation of inorganic nitrate to a cellular nitrogen form. Since this reduction process is essential to phytoplankton growth when nitrate is the only available nitrogen source, the species with the highest capability to reduce nitrate, or the highest nitrate reductase activity, should be the species most favored to dominate a phytoplankton population under nitrate-limiting conditions. To study this hypothesis, the nitrate reductase activities of two species were studied; these species are co-occurring yet dominate under different conditions. Thalassiosira nordenskioldii is the dominant species in Auke Bay, Alaska in early spring when nitrate levels are decreasing from 15 μM to 1 μM, and the temperature is about 5 C. Skeletonema costatum is dominant in mid-summer when nitrate levels are below 1 μM and the temperature is above 12 C. The results show that at 15 C, Skeletonema has a higher enzyme activity at all nitrate levels than does Thalassiosira, which is consistent with the hypothesis. In addition, Thalassiosira shows a higher enzyme activity at 10 C, nearer its optimal temperature for growth, than at 15 C, suggesting that temperature affects species succession through its influence on enzyme activity. The applicability of Michaelis-Menten kinetics to this reduction reaction, based on the enzyme activities measured for these two species, is doubtful but inconclusive. The results also have implications for such concepts as nitrate-limited growth and the Redfield model relating nutrient, O₂ and CO₂ changes in the ocean. / Graduation date: 1971

Recombinant human calpain-5 was expressed in insect cells using a baculovirus system. The expressed calpain-5 was purified by both traditional chromatography and by affinity-column chromatography. Both methods yielded active protease. Calpain-5 displayed very limited hydrophobicity. This indicated that calpain-5 is not a membrane binding protein. Calpain-5 had pI of 8.3. The recombinant calpain-5 also exhibited calcium-dependent proteolytic activity. The calculated calcium requirement for half-maximal activity was 9.6 mM when incubated at 37��C and 26.5 mM when incubated at 30��C. Compared to traditional calpains, which require less than 1 mM calcium for half-maximal activity, calpain-5 exhibited weaker proteolytic activity. This is an unusual observation because calpain-5 lacks the typical calcium-binding domain of the calpains and implied that other calcium-binding region of the protein account for calcium-binding and sensitivity. Our results also showed that calpain-5 was different from traditional calpains because its activity was higher at 37��C compared to 30��C and remained active at 37��C for more than 2 hours. This differs from traditional calpains which display better proteolytic activity at lower temperatures and become inactive within 30 minutes of incubation in 37��C. Calpain-specific inhibitors, calpastatin and E64, did not inhibit calpain-5. Only one calcium-binding inhibitor, PD150606, inhibited calpain-5 proteolytic activity. These results confirmed that calpain's calcium-binding domain is important in calpastatin binding and calpain-5 possesses other calcium-binding regions. Calpain-5 was able to degrade spectrin, a ubiquitous cytoskeletal protein. This indicates that calpain-5 might have a role in cell remodeling. Finally, calpain-5 has the ability to degrade itself. It is not clear if this is the result of inter- or intra-molecular proteolysis and whether this leads to activation of the protein or is, instead, the first step in its degradation. Calpain-5 is expressed at highest concentrations in testis, brain, liver and gastrointestinal tract. It is not clear why these tissues require a unique calpain. Calpain-5 may provide these tissues with an additional calcium-dependent proteolytic activity which is not regulated by calpastatin and which could participate in cytoskeletal protein turnover. / Graduation date: 2004

Graduation date: 1999

Graduation date: 1997