Biochemistry, Biophysics, and Structural Biology Commons^™

Expression Of Cholera Toxin B Subunit-Rotavirus Nsp4 Enterotoxin Fusion Protein In Transgenic Chloroplasts, Anila Kalluri

Electronic Theses and Dissertations

Rotavirus, the major cause of life-threatening infantile gastroenteritis, is a member of the Reoviridae family and is considered to be the single most important cause of virus-based severe diarrheal illness in infants and young children particularly 6 months to 2 years of age in industrialized and developing countries. Infection in infants and young children is often accompanied by severe life threatening diarrhea, most commonly following primary infection. Diarrhea is the major cause of death among children around the world. Responsible for 4 to 6 million deaths per year according to the World Health Organization (WHO), diarrhea is especially dangerous for …

Receptor Mediated Oral Delivery Of Bioencapsulated Green Fluorescent Protein Expressed In Transgenic Chloroplasts, Arati Limaye

Electronic Theses and Dissertations

The skyrocketing costs of prescription medicine in developed countries and their lack of availability in developing countries are the most challenging problems of human health. Primary reasons for such high cost are fermentation-based production, expensive purification methods, the need for low temperature storage and transportation and the delivery through sterile injections. Most of these expenses could be minimized or eliminated when therapeutic proteins are expressed and orally delivered via plant cells. Chloroplasts have the machinery to fold complex and biologically active eukaryotic proteins in the soluble chloroplast stromal compartment. Protein expression through chloroplast transformation system offers a number of advantages …

Correction Of The Literature Citing Monoiodo-8-Quinolinols: A Critical Review / Herman Gershon And Donald D. Clarke Harding Laboratory, The New York Botanical Garde, And Fordham University, Bronx, New York 10458, Usa, Herman Gershon, Donald Dudley Clarke Phd, Donald Dudley Clarke Phd

Chemistry Faculty Publications

Until 1995, all papers that claimed preparation or further study with monoiodo-8-quinolinol were really working with the 7-iodo isomer. Of the 83 relevant citations 67 dealt with incorrect structural assignments, 13 were critiqued due to errors made in synthesis and interpretation, and 9 had the correct structural assignments. This literature review made the following conclusions apparent. 1. Prototropic forms of 8- quinolinol influence orientation of electrophiles. 2. Under strong acidic conditions, the 5 position is favored. 3. Under mild acidic conditions, mixtures of 5 and 7 substituted compounds are formed. 4. Under basic conditions, the incoming electrophile favors the 7 …

Protein Sequence Entropy Is Closely Related To Packing Density And Hydrophobicity, H Liao, W Yeh, D Chiang, R L. Jernigan, Brooke Lustig

Faculty Publications, Chemistry

We investigated the correlation between the Shannon information entropy, ‘sequence entropy’, with respect to the local flexibility of native globular proteins as described by inverse packing density. These are determined at each residue position for a total set of 130 query proteins, where sequence entropies are calculated from each set of aligned residues. For the accompanying aggregate set of 130 alignments, a strong linear correlation is observed between the calculated sequence entropy and the corresponding inverse packing density determined at an associated residue position. This region of linearity spans the range of Cα packing densities from 12 to 25 amino …

Removal Of P-Cresol Sulfate By Hemodialysis, Andres W. Martinez, Natalie S. Recht, Thomas H. Hostetter, Timothy W. Meyer

Chemistry and Biochemistry

Protein-bound solutes are poorly cleared by dialysis. Among the most extensively studied of these solutes is p-cresol, which has been shown to be toxic in vitro. This study examined the form in which p-cresol circulates and quantified its removal by hemodialysis. HPLC analysis of plasma from hemodialysis patients contained a peak whose mobility corresponded to synthetic p-cresol sulfate (PCS) but no detectable unconjugated p-cresol. Treatment with sulfatase resulted in recovery of this peak as p-cresol, confirming its identity. Subsequent studies compared the removal of PCS and another protein-bound solute, indican, to the removal of urea during clinical hemodialysis treatments. …

Selenotrisulfide Derivative Of Alpha-Lipoic Acid: Evaluation In A Cell Culture Model For Potential Use As A Topical Antioxidant, Melenie Lee Alonis

Electronic Theses and Dissertations

Selenium is a required micronutrient in mammalian cells. It is incorporated in the form of selenocysteine into selenoenzymes such as glutathione peroxidase and thioredoxin reductase, and is absolutely required for activity. Thioredoxin reductase is necessary for reduction of oxidized thioredoxin and therefore plays a major role in maintaining the redox status of the cell. Glutathione peroxidase is responsible for reducing peroxides into their corresponding alcohols and water. Together, these selenoenzymes constitute a significant part of the cell's arsenal to defend itself against oxidative stress. Exogenous sources of oxidative stress, such as UV radiation, are capable of generating reactive oxygen species …

Reduced Macrophage Apoptosis Is Associated With Accelerated Atherosclerosis In Low-Denstiy Lipoprotein Receptor-Null Mice, Michael Sinensky, J. Liu, D. P. Thweke, Y. R. Su, M. F. Linton, S. Fazio

Faculty Publications, Biological Sciences

Objective— The majority of apoptotic cells in atherosclerotic lesions are macrophages. However, the pathogenic role of macrophage apoptosis in the development of atherosclerosis remains unclear. Elevated expression of Bax, one of the pivotal proapoptotic proteins of the Bcl-2 family, has been found in human atherosclerotic plaques. Activation of Bax also occurs in free cholesterol-loaded and oxysterol-treated mouse macrophages. In this study, we examined the effect of Bax deficiency in bone marrow-derived leukocytes on the development of atherosclerosis in low-density lipoprotein receptor-null (LDLR−/−) mice. Methods and Results— Fourteen 8-week-old male LDLR−/− mice were lethally irradiated and reconstituted with either wild-type (WT) …

China Lake Buffer Strip Pictures, Colby Environmental Assessment Team, Colby College, Problems In Environmental Science Course (Biology 493), Colby College

Colby College Watershed Study: China Lake (2005, 1989)

No abstract provided.

Ethan's Land Use Photos, Colby Environmental Assessment Team, Colby College, Problems In Environmental Science Course (Biology 493), Colby College

Colby College Watershed Study: China Lake (2005, 1989)

No abstract provided.

Analysis Of Genotypic And Phenotypic Differences In Desmognathus Quadramaculatus Across The Southern Appalachians, Deborah Susan Merritt

Theses, Dissertations and Capstones

Populations of Desmognathus quadramaculatus were compared for genotypic and phenotypic differences across 5 states in the Southern Appalachians. Series consisting of 10 salamanders each were collected from West Virginia, Virginia, Tennessee, North Carolina and Georgia. Salamanders were assessed for genotypic differences by using allozyme (proteins with polymorphic loci) electrophoresis staining for 13 loci of the muscle, liver and stomach tissue. Nei’s genetic distance was used to score genetic variation between populations. Phenotypic differences were assessed by comparing 10 external morphological measurements between populations. Mahalanobis’ distance was used to score morphological differences between populations. Both genetic and morphological differences were correlated …

Computational Study Of A Novel Dinuclear Metal Complex, Dragos Seghete

Inquiry: The University of Arkansas Undergraduate Research Journal

The first compound containing an M-H-M was recently reported by Vicic et al. With recent availability of large computational resources, molecular modeling has become a reliable tool for confirming experimental results. The novel dinuclear Ni complex [( dippm)2Ni2Br2]( 0 -H) was investigated in this work from a theoretical perspective. Full geometry optimization was carried on the dinuclear Ni complex at the DFT/B3LYP with the 6-31G* basis set. The result verifies the linear Ni-H-Ni bond. Two different starting structures that converged to the same geometry confirm that a global minimum was reached. The computed structure differs from the experimentally determined one …

Imposed Constraints On The Smith-Waterman Alignment Algorithm For Enhanced Modeling Of A Single-Molecule Dna Sequencer, Patrick G. Humphrey, Gregory R. Bashford

Biomedical Imaging and Biosignal Analysis Laboratory

An effort has been underway to develop a system for de novo sequencing of single DNA molecules with very long reads. The system operates by optically detecting the passage of fluorescently tagged DNA bases through a detection zone. A successful system would be revolutionary with respect to speed, read length, cost and minimized laboratory infrastructure. An important part of system development is modeling of the detection process. In particular, predicting the expected error from a set of sequencing parameters is helpful in system design. This paper describes variations on the Smith-Waterman algorithm for subsequence alignment used in a single-molecule detection …

Ph-Dependent Substrate Preference Of Pig Heart Lipoamide Dehydrogenase Varies With Oligomeric State: Response To Mitochondrial Matrix Acidification, Natalia L. Klyachko, Valentina A. Shchedrina, Alexander V. Efimov, Sergey V. Kazakov, Irina G. Gazaryan, Bruce S. Kristal, Abraham M. Brown

Department of Biochemistry: Faculty Publications

Cycling of intracellular pH has recently been shown to play a critical role in ischemia-reperfusion injury. Ischemia-reperfusion also leads to mitochondrial matrix acidification and dysfunction. However, the mechanism by which matrix acidification contributes to mitochondrial dysfunction, oxidative stress, and the resultant cellular injury has not been elucidated. We observe pH-dependent equilibria between monomeric, dimeric, and a previously undescribed tetrameric form of pig heart lipoamide dehydrogenase (LADH), a mitochondrial matrix enzyme. Dynamic light scattering studies of native LADH in aqueous solution indicate that lowering pH favors a shift in average molecular mass from higher oligomeric states to monomer. Sedimentation velocity of …

An Algorithm For Identification Of Bacterial Selenocysteine Insertion Sequence Elements And Selenoprotein Genes, Yan Zhang, Vadim Gladyshev

Department of Biochemistry: Faculty Publications

Motivation: Incorporation of selenocysteine (Sec) into proteins in response to UGA codons requires a cis-acting RNA structure, Sec insertion sequence (SECIS) element. Whereas SECIS elements in Escherichia coli are well characterized, a bacterial SECIS consensus structure is lacking.

Results: We developed a bacterial SECIS consensus model, the key feature of which is a conserved guanosine in a small apical loop of the properly positioned structure. This consensus was used to build a computational tool, bSECISearch, for detection of bacterial SECIS elements and selenoprotein genes in sequence databases. The program identified 96.5% of known selenoprotein genes in completely sequenced bacterial genomes …

Regulation Of Enteric Endophytic Bacterial Colonization By Plant Defenses, A. Leonardo Iniguez, Yuemei Dong, Heather D. Carter, Brian M. M. Ahmer, Julie M. Stone, Eric W. Triplett

Department of Biochemistry: Faculty Publications

Bacterial endophytes reside within the interior of plants without causing disease or forming symbiotic structures. Some endophytes, such as Klebsiella pneumoniae 342 (Kp342), enhance plant growth and nutrition. Others, such as Salmonella enterica serovar Typhimurium (S. typhimurium), are human pathogens that contaminate raw produce. Several lines of evidence are presented here to support the hypothesis that plant defense response pathways regulate colonization by endophytic bacteria. An ethyleneinsensitive mutant of Medicago truncatula is hypercolonized by Kp342 compared to the parent genotype. Addition of ethylene, a signal molecule for induced systemic resistance in plants, decreased endophytic colonization in Medicago spp. …

Diversity And Functional Plasticity Of Eukaryotic Selenoproteins: Identification And Characterization Of The Selj Family, Sergi Castellano, Alexey V. Lobanov, Charles Chapple, Sergey V. Novoselov, Mario Albrecht, Deame Hua, Alain Lescure, Thomas Lengauer, Alain Krol, Vadim N. Gladyshev, Roderic Guigó

Department of Biochemistry: Faculty Publications

Selenoproteins are a diverse group of proteins that contain selenocysteine (Sec), the 21st amino acid. In the genetic code, UGA serves as a termination signal and a Sec codon. This dual role has precluded the automatic annotation of selenoproteins. Recent advances in the computational identification of selenoprotein genes have provided a first glimpse of the size, functions, and phylogenetic diversity of eukaryotic selenoproteomes. Here, we describe the identification of a selenoprotein family named SelJ. In contrast to known selenoproteins, SelJ appears to be restricted to actinopterygian fishes and sea urchin, with Cys homologues only found in cnidarians. SelJ shows significant …

Phylogenetic Engineering At An Interface Between Large And Small Subunits Imparts Land-Plant Kinetic Properties To Algal Rubisco, Robert J. Spreitzer, Srinivasa R. Peddi, Sriram Satagopan

Department of Biochemistry: Faculty Publications

Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) catalyzes the rate-limiting step of photosynthetic CO₂ fixation and, thus, limits agricultural productivity. However, Rubisco enzymes from different species have different catalytic constants. If the structural basis for such differences were known, a rationale could be developed for genetically engineering an improved enzyme. Residues at the bottom of the large-subunit α/β-barrel active site of Rubisco from the green alga Chlamydomonas reinhardtii (methyl-Cys-256, Lys-258, and Ile-265) were previously changed through directed mutagenesis and chloroplast transformation to residues characteristic of land-plant Rubisco (Phe-256, Arg-258, and Val-265). The resultant enzyme has decreases in carboxylation efficiency and CO₂/O …

Condensed Protocol For Competent Cell Preparation And Transformation Of The Methylotrophic Yeast Pichia Pastoris, Joan Lin-Cereghino, William Wong, See Xiong, William Giang, Linda Luong, Jane Vu, Sabrina Johnson, Geoff P. Lin-Cereghino

College of the Pacific Faculty Articles

No abstract provided.

Biphasic Functions Of The Kinase-Defective Ephb6 Receptor In Cell Adhesion And Migration, Hiroshi Matsuoka, Hiroya Obama, Meghan Kelly, Toshimitsu Matsui, Masaru Nakamoto

Biology Faculty Publications

EphB6 is a unique member in the Eph family of receptor tyrosine kinases in that its kinase domain contains several alterations in conserved amino acids and is catalytically inactive. Although EphB6 is expressed both in a variety of embryonic and adult tissues, biological functions of this receptor are largely unknown. In the present study, we examined the function of EphB6 in cell adhesion and migration. We demonstrated that EphB6 exerted biphasic effects in response to different concentrations of the ephrin-B2 ligand; EphB6 promoted cell adhesion and migration when stimulated with low concentrations of ephrin-B2, whereas it induced repulsion and inhibited …

Selective Field Effects On Intracellular Vacuoles And Vesicle Membranes With Nanosecond Electric Pulses, Ephrem Tekle, Hammou Oubrahim, Sergey M. Dzekunov, Juergen F. Kolb, Karl H. Schoenbach

Bioelectrics Publications

Electric pulses across intact vesicles and cells can lead to transient increase in permeability of their membranes. We studied the integrity of these membranes in response to external electric pulses of high amplitude and submicrosecond duration with a primary aim of achieving selective permeabilization. These effects were examined in two separate model systems comprising of 1), a mixed population of 1,2-di-oleoyl-sn-glycero-3-phosphocholine phospholipid vesicles and in 2), single COS-7 cells, in which large endosomal membrane vacuoles were induced by stimulated endocytosis. It has been shown that large and rapidly varying external electric fields, with pulses shorter than the charging time of …

Computer-Assisted Docking Of Flavodoxin With The Atp:Co(I)Rrinoid Adenosyltransferase (Coba) Enzyme Reveals Residues Critical For Protein-Protein Interactions But Not For Catalysis, Nicole R. Baun, Jorge C. Escalante-Semerena

Department of Biochemistry: Faculty Publications

The activity of the housekeeping ATP:co(I)rrinoid adenosyltransferase

(CobA) enzyme of Salmonella enterica sv. Typhimurium

is required to adenosylate de novo biosynthetic intermediates

of adenosylcobalamin and to salvage incomplete and complete

corrinoids from the environment of this bacterium. In vitro,

reduced flavodoxin (FldA) provides an electron to generate the

co(I)rrinoid substrate in the CobA active site. To understand how

CobAand FldA interact, a computer model of aCobA-FldA complex

was generated. This model was used to guide the introduction of

mutations into CobA using site-directed mutagenesis and the synthesis

of a peptide mimic of FldA. Residues Arg-9 and Arg-165 of

CobA …

The Effects Of Β-Mercaptoethanol On The Specific Activity Of Porphobilinogen Synthase Mutants Of The Rhodobacter Sphaeroides Protein, Trefan Archibald '05

Honors Projects

The enzyme porphobilinogen synthase (PBGS) catalyzes the conversion of two molecules of δ-aminolevulinic acid (ALA) into porphobilinogen (PBG) in the first common step of the tetrapyrrole biosynthesis pathway. A homology model of the R. sphaeroides PBGS was created by Dr. Eileen K. Jaffe at Fox Chase Cancer Center based on comparison to the crystal structure of Pseudomonas aeruginosa. The proposed structure suggests that there are four cysteines in close proximity to the active site. Three of these cysteines are not present in the highly similar R. capsulatus sequence. Under oxidizing conditions these residues can potentially participate in the formation of …

Host Blood Proteins And Peptides In The Midgut Of The Tick Dermacentor Variabilis Contribute To Bacterial Control, Daniel E. Sonenshine, Wayne L. Hynes, Shane M. Ceraul, Robert Mitchell, Tiffany Benzine

Biological Sciences Faculty Publications

Antimicrobial midgut proteins and peptides that result from blood digestion in feeding American dog ticks Dermacentor variabilis (Say) were identified. Midgut extracts from these ticks showed antimicrobial activity against Micrococcus luteus, regardless of whether they were challenged with peptidoglycan, blood meal components, rabbit blood, Bacillus subtilis, Escherischia coli or Borrelia burgdorferi. However, no peptide band co-migrating with defensin was found in midgut extracts from the challenged ticks. Partial purification of the midgut extracts using C₁₈ Sep Paks and gel electrophoresis showed the presence of 4 distinct bands with rMW 4.1, 5.3, 5.7 and 8.0 kDa identified …

Effect Of Humic Substance Photodegradation On Bacterial Growth And Respiration In Lake Water, Alexandre M. Anesio, William Granéli, George R. Aiken, David J. Kieber, Kenneth Mopper

Chemistry & Biochemistry Faculty Publications

This study addresses how humic substance (HS) chemical composition and photoreactivity affect bacterial growth, respiration, and growth efficiency (BGE) in lake water. Aqueous solutions of HSs from diverse aquatic environments representing different dissolved organic matter sources (autochthonous and allochthonous) were exposed to artificial solar UV radiation. These solutions were added to lake water passed through a 0.7-μm pore-size filter (containing grazer-free lake bacteria) followed by dark incubation for 5, 43, and 65 h. For the 5-h incubation, several irradiated HSs inhibited bacterial carbon production (BCP) and this inhibition was highly correlated with H₂O₂ photoproduction. The H_{2 …}

Cytokine And Chemokine Profiles In A Rat Model Of Hemorrhagic Shock After Immuno-Modulation By Androstenetriol, Kristin E. Paccione

Theses and Dissertations

Further understanding of the cellular and molecular mechanisms involved in traumatic injury, and how they are modulated during drug interaction, can facilitate novel treatment strategies for future trauma patients. We hypothesize that the pharmacological agent, Androstenetriol (AET), up regulates host immune response by modulating the continued expression of mediators, including cytokines. In a double-blinded experiment, rats were hemorrhaged, driven by volume or pressure conditions, then resuscitated with fluids and packed red blood cells following a subcutaneous injection of either vehicle or drug. Blood was collected at various time points and cytokine levels were determined by analyses with both multiplex and …

Identification Of The Rna Cis-Elements That Interact With Srp30a To Regulate The Alternative Splicing Of Caspase 9 Pre-Mrna, Prabhat Mukerjee

Theses and Dissertations

Studies have shown that the alternative splicing of caspase 9 and the phospho-status of SR proteins, a conserved family of splicing factors, are regulated by chemotherapy and de novo ceramide via the action of protein phosphatase-1 (PP1). Two RNA splice variants are derived from the caspase 9 gene, pro-apoptotic caspase 9a and anti-apoptotic caspase 9b, via alternative splicing by either the inclusion or exclusion of an exon 3, 4, 5, and 6 cassette. In this study, the link between SR proteins and the alternative splicing of caspase 9 was established. Sequence analysis of the exon 3, 4, 5, and 6 …

Mapping Of Ssdna Nicks Within Dsdna Genomes By Two-Dimensional Gel Electrophoresis, Wayne P. Wahls, K. Mark Dewall, Mari K. Davidson

Journal of the Arkansas Academy of Science

DNA molecules within chromosomes undergo constant, dynamic changes yet maintain the integrity of the primary DNA sequence. DNA replication, adjustment of helical density, resolution of catenenes, repair of DNA damage, and homologous recombination each involve breakage and religation of the phosphate backbone of the double helix. Although the analysis of dsDNA breaks is facile, the analysis of ssDNA nicks is not. The principal impediment is that conventional, one-dimensional electrophoresis methods cannot readily detect ssDNA nicks in the context of dsDNA breaks. We therefore developed a two dimensional (native/denaturing) gel electrophoresis approach to map the positions of ssDNA nicks. Analysis of …

Arsenite Regulates Cystic Fibrosis Transmem­Brane Conductance Regulator And P-Glycoprotein: Evidence Of Pathway Independence, Rangan Maitra, Joshua Hamilton

Dartmouth Scholarship

In the past, people have argued for and against the theory of reciprocal regulation of the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) and P-glycoprotein (Pgp). Data have indicated that this may occur in vitro during drug-induced selection of cells, and in vivo during development. Much of this debate has been caused by a severe lack of mechanistic details involved in such regulation. Our past data indicate that certain Pgp modulators can affect CFTR expression and function. The goal of this study was to investigate the effects of trivalent arsenic (arsenite), a known transcriptional activator of Pgp, on CFTR expression. In …

A Novel Binding Interaction For The Paxillin Ld3 Motif: Paxillin Ld3 Mediates Merlin-Paxillin Binding At Paxillin Binding Domain 1, Sandra E. Beattie Geden

Electronic Theses and Dissertations

Neurofibromatosis type 2, an autosomal dominant genetic disorder, causes predisposed individuals to develop various benign central and peripheral nervous system tumors. The characteristic tumors of this disease are schwannomas, which are tumors of the Schwann cells, typically on the vestibular nerve. These and the other associated tumors slowly compress nervous system structures causing deafness and loss of balance, resulting in an average life-span of less than 40 years. The product of the Nf2 gene is the protein named merlin or schwannomin. In individuals diagnosed with NF2, merlin is either absent or mutated to the point of inactivation. As such, merlin …

Modulation Of Gene Expression By Tumor-Derived Mutant P53. Role Of Transactivation In Gain-Of-Function., Mariano J. Scian

Theses and Dissertations

It was hypothesized that the C-terminal sequences for mutant p53 would be required for oligomerization. and oligomerization may be critical for gain-of-function. An N-terminal deletion mutant of p53 that deletes amino acids 1-293 was used as a tool to perform hetero-oligomerization studies. This mutant retains the entire oligomerization domain but dispenses off the transactivation domain and a large portion of the sequence- specific DNA-binding domain. Co-transfection experiments show that p53 del. 1-293 forms hetero-oligomeric complexes with p53-D281G. Also. co-expression of p53 del. 1- 293 with p53-D281G inhibited p53-D28lG-mediated transactivation of the EGFR and MDRl promoters suggesting that hetero-oligomerization inactivates transcriptional …