Verlags-Signet [Illustration] : (der Firma S. Calvary & Co. zum 50jährigen Jubiläum - Mai 1902 - von ihrem Personal gewidmet)

Hermann Struck

Lesser Ury-Ausstellung im Salon von Keller & Reiner

M. Rapsilber

Jean Richard Bloch, Simler & Co. [Rezension]

S.

Measurement of Protein in Nearshore Marine Sediments

Proteinaceous material in marine sediments which is available to proteolytic hydrolysis has been measured using a new method. This technique utilizes Coomassie Blue dye binding, which has the advantage of being sensitive only to larger polypeptides. Substantial interferences from other sedmentary organic substances are overcome by using a standard additions approach in conjunction with enzymatic digestion of the protein. Although tedious, the technique provides acceptable precision and accuracy. Measurements of protein in surficial nearshore sediments of the Gulf of Maine and St. Croix yield values ranging from 0.1 to 2.2 mg g-1, which account for a minor fraction of total nitrogen or acid-hydrolyzable amino acids. Protein decreases downcore at a faster rate than either of these 2 indicators of nitrogenous material, indicating the greater lability of the truly proteinaceous material. Biomass comprises a minor portion of the measured protein.

Animal Guts as Nonideal Chemical Reactors: Partial Mixing and Axial Variation in Absorption Kinetics

Animal guts have been idealized as axially uniform plug-flow reactors (PFRs) without significant axial mixing or as combinations in series of such PFRs with other reactor types. To relax these often unrealistic assumptions and to provide a means for relaxing others, I approximated an animal gut as a series of n continuously stirred tank reactors (CSTRs) and examined its performance as a Function of n. For the digestion problem of hydrolysis and absorption in series, I suggest as a first approximation that a tubular gut of length L and diameter D comprises n=L/D tanks in series. For n greater than or equal to 10, there is little difference between performance of the nCSTR model and an ideal PFR in the coupled tasks of hydrolysis and absorption. Relatively thinner and longer guts, characteristic of animals feeding on poorer forage, prove more efficient in both conversion and absorption by restricting axial mixing, in the same total volume, they also give a higher rate of absorption. I then asked how a fixed number of absorptive sites should be distributed among the n compartments. Absorption rate generally is maximized when absorbers are concentrated in the hindmost few compartments, but high food quality or suboptimal ingestion rates decrease the advantage of highly concentrated absorbers. This modeling approach connects gut function and structure at multiple scales and can be extended to include other nonideal reactor behaviors observed in real animals.

Production of Ethanol from Hardwood

Diminishing crude oil and natural gas supplies, along with concern about greenhouse gas are major driving forces in the search for efficient renewable energy sources. The conversion of lignocellulosic biomass to energy and useful chemicals is a component of the solution. Ethanol is most commonly produced by enzymatic hydrolysis of complex carbohydrates to simple sugars followed by fermentation using yeast. C6Hl0O5 + H2O −Enxymes→ C6H12O6 −Yeast→ 2CH3CH2OH + 2C02 In the U.S. corn is the primary starting raw material for commercial ethanol production. However, there is insufficient corn available to meet the future demand for ethanol as a gasoline additive. Consequently a variety of processes are being developed for producing ethanol from biomass; among which is the NREL process for the production of ethanol from white hardwood. The objective of the thesis reported here was to perform a technical economic analysis of the hardwood to ethanol process. In this analysis a Greenfield plant was compared to co-locating the ethanol plant adjacent to a Kraft pulp mill. The advantage of the latter case is that facilities can be shared jointly for ethanol production and for the production of pulp. Preliminary process designs were performed for three cases; a base case size of 2205 dry tons/day of hardwood (52 million gallons of ethanol per year) as well as the two cases of half and double this size. The thermal efficiency of the NREL process was estimated to be approximately 36%; that is about 36% of the thermal energy in the wood is retained in the product ethanol and by-product electrical energy. The discounted cash flow rate of return on investment and the net present value methods of evaluating process alternatives were used to evaluate the economic feasibility of the NREL process. The minimum acceptable discounted cash flow rate of return after taxes was assumed to be 10%. In all of the process alternatives investigated, the dominant cost factors are the capital recovery charges and the cost of wood. The Greenfield NREL process is not economically viable with the cost of producing ethanol varying from $2.58 to $2.08/gallon for the half capacity and double capacity cases respectively. The co-location cases appear more promising due to reductions in capital costs. The most profitable co-location case resulted in a discounted cash flow rate of return improving from 8.5% for the half capacity case to 20.3% for the double capacity case. Due to economy of scale, the investments become more and more profitable as the size of the plant increases. This concept is limited by the amount of wood that can be delivered to the plant on a sustainable basis as well as the demand for ethanol within a reasonable distance of the plant.

Kinetics and Mechanism of Oxygen Delignification

Considerable research has been conducted into the kinetics and selectivity of the oxygen delignification process to overcome limitation in its use. However most studies were performed in a batch reactor whereby the hydroxide and dissolved oxygen concentrations are changing during the reaction time in an effort to simulate tower performance in pulp mills. This makes it difficult to determine the reaction order of the different reactants in the rate expressions. Also the lignin content and cellulose degradation of the pulp are only established at the end of the experiment when the sample is removed from the batch reactor. To overcome these deficiencies, we have adopted a differential reactor system used frequently for fluid-solid rate studies (so-called Berty reactor) for measurement of oxygen delignification kinetics. In this reactor, the dissolved oxygen concentration and the alkali concentration in the feed are kept constant, and the rate of lignin removal is determined from the dissolved lignin content in the outflow stream measured by UV absorption. The mass of lignin removed is verified by analyzing the pulp at several time intervals. Experiments were performed at different temperatures, oxygen pressures and caustic concentrations. The delignification rate was found to be first order in HexA-free residual lignin content. The delignification rate reaction order in caustic concentration and oxygen pressure were determined to be 0.42 and 0.44 respectively. The activation energy was found to be 53kJ/mol. The carbohydrate degradation during oxygen delignification can be described by two contributions: one due to radicals produced by phenolic delignification, and a much smaller contribution due to alkaline hydrolysis. From the first order of the reaction and the pKa of the active lignin site, a new oxygen delignification mechanism is proposed. The number 3 carbon atom in the aromatic ring with the attached methoxyl group forms the lignin active site for oxygen adsorption and subsequent electrophic reaction to form a hydroperoxide with a pKa value similar to that of the present delignification kinetics. The uniform presence of the aromatic methoxyl groups in residual lignin further support the first order in lignin kinetics.

VERIFICATION OF DNA PREDICTED PROTEIN SEQUENCES BY ENZYME HYDROLYSIS AND MASS SPECTROMETRIC ANALYSIS

The focus of this thesis lies in the development of a sensitive method for the analysis of protein primary structure which can be easily used to confirm the DNA sequence of a protein's gene and determine the modifications which are made after translation. This technique involves the use of dipeptidyl aminopeptidase (DAP) and dipeptidyl carboxypeptidase (DCP) to hydrolyze the protein and the mass spectrometric analysis of the dipeptide products.^ Dipeptidyl carboxypeptidase was purified from human lung tissue and characterized with respect to its proteolytic activity. The results showed that the enzyme has a relatively unrestricted specificity, making it useful for the analysis of the C-terminal of proteins. Most of the dipeptide products were identified using gas chromatography/mass spectrometry (GC/MS). In order to analyze the peptides not hydrolyzed by DCP and DAP, as well as the dipeptides not identified by GC/MS, a FAB ion source was installed on a quadrupole mass spectrometer and its performance evaluated with a variety of compounds.^ Using these techniques, the sequences of the N-terminal and C-terminal regions and seven fragments of bacteriophage P22 tail protein have been verified. All of the dipeptides identified in these analysis were in the same DNA reading frame, thus ruling out the possibility of a single base being inserted or deleted from the DNA sequence. The verification of small sequences throughout the protein sequence also indicates that no large portions of the protein have been removed after translation. ^

THE EFFECT OF RELAXIN ON BIOCHEMICAL AND MORPHOLOGICAL PARAMETERS OF RAT MYOMETRIAL CELLS IN CULTURE (CYCLIC-AMP, CELL SHAPE CHANGE)

Relaxin is able to inhibit spontaneous, oxytocin-and prostaglandin-driven uterine contractions. The intracellular mechanism of action of relaxin on uterine relaxation had previously been studied using isometrically suspended uterine strips. Since uterine strips contain stroma as well as myometrium, the changes in biochemical parameters induced by relaxin treatment may not occur in the same cell types responsible for the physical changes. In these studies, cultures of enriched populations of rat myometrial cells were used to investigate the effect of relaxin on biochemical and morphological parameters which are related to relaxation.^ Under optimal culture conditions (initial plating density 1 - 1.5 x 10('6)cells/ml, 3 ml/35 mm dish, 2 days culture), enzymatically isolated rat myometrial cells were able to respond to relaxin with cAMP elevation. Relaxin elevated cAMP levels in the presence but not the absence of 0.1 mM methylisobutylxanthine or 0.4 um forskolin in a time- and concentration-dependent manner. In contrast, isoproterenol was able to elevate cAMP levels in the presence and absence of 0.1 mM methylisobutylxanthine.^ Oxytocin treatment caused a decrease in mean cell length and area of myometrial cells in culture which could be considered analogous to contraction. Under optimal culture conditions, relaxin increased myometrial cell length and area (i.e. analogous to relaxation) of oxytocin-treated cells in a time- and concentration-dependent manner. Other relaxants such as isoproterenol and dibutyryl cAMP also increased cell length and area of oxytocin - treated myometrial cells in culture.^ Under optimal culture conditions, relaxin decreased myosin light chain kinase activity in a time-and concentration-dependent manner by increasing the K(,50) of the enzyme for calmodulin (CaM), i.e. decreasing the affinity of the enzyme for CaM. The decrease in the affinity of myosin light chain kinase for CaM may be due to the phosphorylation of the enzyme by cAMP-dependent protein kinase. Relaxin also decreased the Ca('2+)(.)CaM-independent myosin light chain kinase activity to a lesser extent than that of the Ca('2+)(.)CaM-dependent enzyme activity. This was not attributable to a decrease in the affinity of the enzyme for myosin in myometrial cells in culture, in contrast to the finding of such a change following relaxin treatment of uterine strips. Further studies are required to clarify this point.^ There was a temporal association between the effects of relaxin on elevation of cAMP levels in the presence of 0.4 uM forskolin, increase in cell length and decrease in myosin light chain kinase activity. . . . (Author's abstract exceeds stipulated maximum length. Discontinued here with permission of author.) UMI ^

REGULATION OF BRAIN NORADRENERGIC-RECEPTOR FUNCTION BY ADRENOCORTICOTROPHIN AND ANTIDEPRESSANT DRUGS (ADRENERGIC RECEPTOR, CYCLIC-AMP, ADENYLATE CYCLASE, IMIPRAMINE, STRESS)

Human behavior appears to be regulated in part by noradrenergic transmission since antidepressant drugs modify the number and function of (beta)-adrenergic receptors in the central nervous system. Affective illness is also known to be associated with the endocrine system, particularly the hypothalamic-pituitary-adrenal axis. The aim of the present study was to determine whether hormones, in particular adrencorticotrophin (ACTH) and corticosterone, may influence behavior by regulating brain noradrenergic receptor function.^ Chronic treatment with ACTH accelerated the increase or decrease in rat brain (beta)-adrenergic receptor number induced by a lesion of the dorsal noradrenergic bundle or treatment with the antidepressant imipramine. Chronic administration of ACTH alone had no effect on (beta)-receptor number although it reduced norepinephrine stimulated cyclic AMP accumulation in brain slices. Treatment with imipramine also reduced the cyclic AMP response to norepinephrine but was accompanied by a decrease in (beta)-adrenergic receptor number. Both the imipramine and ACTH treatments reduced the affinity of (beta)-adrenergic receptors for norepinephrine, but only the antidepressant modified the potency of the neurotransmitter to stimulate second messenger production. Neither ACTH nor imipramine treatment altered Gpp(NH)p- or fluoride-stimulated adenylate cyclase, cyclic AMP, cyclic GMP, or cyclic GMP-stimulated cyclic AMP phosphodiesterase, or the activity of the guanine nucleotide binding protein (Gs). These findings suggested that post-receptor components of the cyclic nucleotide generating system are not influenced by the hormone or antidepressant. This conclusion was verified by the finding that neither treatment altered adenosine-stimulated cyclic AMP accumulation in brain tissue.^ A detailed examination of the (alpha)- and (beta)-adrenergic receptor components of norepinephrine-stimulated cyclic AMP production revealed that ACTH, but not imipramine, administration reduced the contribution of the (alpha)-receptor mediated response. Like ACTH treatment, corticosterone diminished the (alpha)-adrenergic component indicating that adrenal steroids probably mediate the neurochemical responses to ACTH administration. The data indicate that adrenal steroids and antidepressants decrease noradrenergic receptor function by selectively modifying the (alpha)- and (beta)-receptor components. The functional similarity in the action of the steroid and antidepressants suggests that adrenal hormones normally contribute to the maintenance of receptor systems which regulate affective behavior in man. ^

Orléans-Guérin-Porges & Co.

A. G. von Suttner

Regulation of Protein Degradation in the Heart by AMP-Activated Protein Kinase

The degradation of proteins by the ubiquitin proteasome system is essential for cellular homeostasis in the heart. An important regulator of metabolic homeostasis is AMP-activated protein kinase (AMPK). During nutrient deprivation, AMPK is activated and intracellular proteolysis is enhanced through the ubiquitin proteasome system (UPS). Whether AMPK plays a role in protein degradation through the UPS in the heart is not known. Here I present data in support of the hypothesis that AMPK transcriptionally regulates key players in the UPS, which, under extreme conditions can be detrimental to the heart. The ubiquitin ligases MAFbx /Atrogin-1 and MuRF1, key regulators of protein degradation, and AMPK activity are increased during nutrient deprivation. Pharmacologic and genetic activation of AMPK is sufficient for the induction of MAFbx/Atrogin-1 and MuRF1 in cardiomyocytes and in the heart in vivo. Comprehensive experiments demonstrate that the molecular mechanism by which AMPK regulates MuRF1 expression is through the transcription factor myocyte enhancer factor 2 (MEF2), which is involved in stress response and cardiomyocyte remodeling. MuRF1 is required for AMPK-mediated protein degradation through the UPS in cardiomyocytes. Consequently, the absence of MuRF1 during chronic fasting preserves cardiac function, possibly by limiting degradation of critical metabolic enzymes. Furthermore, during cardiac hypertrophy, chronic activation of AMPK also leads to cardiac dysfunction, possibly through enhanced protein degradation and metabolic dysregulation. Collectively, my findings demonstrate that AMPK regulates expression of ubiquitin ligases which are required for UPS-mediated protein degradation in the heart. Based on these results, I propose that specific metabolic signals may serve as modulators of intracellular protein degradation in the heart.

Regulation of Protein Degradation in the Heart by AMP-Activated Protein Kinase

The degradation of proteins by the ubiquitin proteasome system is essential for cellular homeostasis in the heart. An important regulator of metabolic homeostasis is AMP-activated protein kinase (AMPK). During nutrient deprivation, AMPK is activated and intracellular proteolysis is enhanced through the ubiquitin proteasome system (UPS). Whether AMPK plays a role in protein degradation through the UPS in the heart is not known. Here I present data in support of the hypothesis that AMPK transcriptionally regulates key players in the UPS, which, under extreme conditions can be detrimental to the heart. The ubiquitin ligases MAFbx /Atrogin-1 and MuRF1, key regulators of protein degradation, and AMPK activity are increased during nutrient deprivation. Pharmacologic and genetic activation of AMPK is sufficient for the induction of MAFbx/Atrogin-1 and MuRF1 in cardiomyocytes and in the heart in vivo. Comprehensive experiments demonstrate that the molecular mechanism by which AMPK regulates MuRF1 expression is through the transcription factor myocyte enhancer factor 2 (MEF2), which is involved in stress response and cardiomyocyte remodeling. MuRF1 is required for AMPK-mediated protein degradation through the UPS in cardiomyocytes. Consequently, the absence of MuRF1 during chronic fasting preserves cardiac function, possibly by limiting degradation of critical metabolic enzymes. Furthermore, during cardiac hypertrophy, chronic activation of AMPK also leads to cardiac dysfunction, possibly through enhanced protein degradation and metabolic dysregulation. Collectively, my findings demonstrate that AMPK regulates expression of ubiquitin ligases which are required for UPS-mediated protein degradation in the heart. Based on these results, I propose that specific metabolic signals may serve as modulators of intracellular protein degradation in the heart.

Proceedings of the 13th Annual Biochemical Engineering Symposium

The symposium reported here was the thirteenth of a series devoted to talks by students on their biochemical engineering research. The first, third, fifth, ninth, and twelfth were at Kansas State University, the second and fourth were at the University of Nebraska–Lincoln, the sixth was in Kansas City and was hosted by Iowa State University, the seventh and tenth were at Iowa State, and the eighth and eleventh were at the University of Missouri–Columbia and Colorado State University, respectively. All symposia have been followed by proceedings edited by faculty of the host institution. Because final publication usually takes place elsewhere, papers here are brief, and often cover research in progress. ContentSequential Utilization of Mixed Sugars by Clostridium acetobutylicum, B. Hong, N. H. Choi, and L. T. Fan, Kansas State University The Effects of Dilution Rate on the Kinetics. of Anaerobic Acidogenesis, C. J. Huang, Colorado State University Ethanol Production by Zymomonas mobilis in Anaerobic Glucose-Limited Culture: A Yield Study, Mehmet D. Oner, Kansas State University Hydrolysis of Cellulosics by Enterobacteria, Michael R. Sierks, Iowa State University The Cellulase System of Chaetomium cellulolyticum, Nikhil Mehta, Colorado State University DNA Measurement as a Tool for Estimating Biomass Concentration in the Presence of Interfering Solids, Bamidele 0. Solomon, Kansas State University The Effect of Cellulose Crystallinity on Enzymatic Hydrolysis, Maria S. Bertran, Colorado State University High Performance Liquid Chromatography of Di- and Trisaccharides, Michael M. Meagher, Iowa State University Dynamics of Bubble Size .Distributions in Air-Lift Fermentors, c. H. Lee and Snehal A. Patel, Kansas State University A Thermal Coagulation Study of Alfalfa Leaf Proteins by Differential Scanning Calorimeter, Khalif Ahmed and Bruce Dale, Colorado State University Thermodynamic Efficiency of Photoautotrophic Growth, Hyeon Y. Lee, Kansas State University

Proceedings of the 14th Annual Biochemical Engineering Symposium

The Annual Biochemical Engineering Symposium series is devoted to presentations by students on their research topics. The fourteenth event, held in 1984, was organized at the University of Missouri–Columbia. It was attended by the biochemical engineering faculty and the students from Colorado State University, Iowa State University, Kansas State University, University of Missouri–Columbia, University of Missouri–Rolla and Washington University, St. Louis. Contents"Estimation of Product Formation Kinetics and Microbial Yield Parameters for Anaerobic Organic Acid and Solvent Production," M.D. Oner, Kansas State University "Characterization of Soy Protein Texturization in a Complex Bioreactor," J.L. Ibave, Colorado State University "Acid and Solvent Fermentations Using Mixed Cultures," D. Stevens, University of Missouri–Columbia "Preliminary Process Design for Ethanol from Sweet Sorghum Ensilage Feedstock," Keith D. Lange, Colorado State University "Lamella Settlers in Ethanol Fermentation," Yong Jayanata, University of Missouri–Columbia "Bubble Size Distribution in the Down Flow Section of an Air-Lift Column," Snehal A. Patel and C.H. Lee, Kansas State University "The Sensitivity of Plant Cells to Shear Stress," Morris Z. Resenberg and Eric H. Dunlap, Washington University, St. Louis "Estimation of Growth Yield Parameters Associated with Microbial Growth," Hyeon Y. Lee, Kansas State University "Capillary Gas Chromatography of Trimethylsilylated Trisaccharides," Etienne J.M. Selosse, Iowa State University "Subsite Mapping of an Endo-Xylanase Labeled Xylooligo-saccharides," Bernard Y. Tao, Iowa State University "Cellulase Enzyme Recycle," Kate M.V. Baptie, Colorado State University "Non-Homogeneous Poisson Renewal Reward Process for Modelling Enzymatic Hydrolysis of Cellulose," M.M. Gharpuray and L.T. Fan, Kansas State University