Biochemistry Commons^™

Identification Of Disufide Bond Formation Between Mitoneet And Glutamate Dehydrogenase 1, Morgan E. Roberts, Jacquelyn P. Crail, Megan M. Laffoon, William G. Fernandez, Michael A. Menze, Mary E. Konkle

Michael Menze

MitoNEET is a protein that was identified as a drug target for diabetes, but its cellular function as well as its role in diabetes remains elusive. Protein pull-down experiments identified glutamate dehydrogenase 1 (GDH1) as a potential binding partner. GDH1 is a key metabolic enzyme with emerging roles in insulin regulation. MitoNEET forms a covalent complex with GDH1 through disulfide bond formation and acts as an activator. Proteomic analysis identified the specific cysteine residues that participate in the disulfide bond. This is the first report that effectively links mitoNEET to activation of the insulin regulator GDH1.

Interplay Of Hydrogen Bonds And N→Π* Interactions In Proteins, Gail J. Bartlett, Robert W. Newberry, Brett Vanveller, Ronald T. Raines, Derek N. Woolfson

Brett VanVeller

Protein structures are stabilized by multiple weak interactions, including the hydrophobic effect, hydrogen bonds, electrostatic effects, and van der Waals interactions. Among these interactions, the hydrogen bond is distinct in having its origins in electron delocalization. Recently, another type of electron delocalization, the n→π* interaction between carbonyl groups, has been shown to play a role in stabilizing protein structure. Here we examine the interplay between hydrogen bonding and n→π* interactions. To address this issue, we used data available from high-resolution protein crystal structures to interrogate asparagine side-chain oxygen atoms that are both acceptors of a hydrogen bond and donors of …

The Interdomain Interface In Bifunctional Enzyme Protein 3/4a (Ns3/4a) Regulates Protease And Helicase Activities, Cihan Aydin, Sourav Mukherjee, Alicia Hanson, David Frick, Celia Schiffer

Celia A. Schiffer

Hepatitis C (HCV) protein 3/4A (NS3/4A) is a bifunctional enzyme comprising two separate domains with protease and helicase activities, which are essential for viral propagation. Both domains are stable and have enzymatic activity separately, and the relevance and implications of having protease and helicase together as a single protein remains to be explored. Altered in vitro activities of isolated domains compared with the full-length NS3/4A protein suggest the existence of interdomain communication. The molecular mechanism and extent of this communication was investigated by probing the domain-domain interface observed in HCV NS3/4A crystal structures. We found in molecular dynamics simulations that …

Detection Of Boronic Acids Through Excited-State Intramolecular Proton-Transfer Fluorescence, Matthew R. Aronoff, Brett Vanveller, Ronald T. Raines

Brett VanVeller

Boronic acids are versatile reagents for the chemical synthesis of organic molecules. They and other boron-containing compounds can be detected readily by the interruption of the excited-state intramolecular proton transfer (ESIPT) of 10-hydroxybenzo[h]quinolone. This method is highly sensitive and selective, and useful for monitoring synthetic reactions and detecting boron-containing compounds on a solid support.

A Divalent Protecting Group For Benzoxaboroles, Brett Vanveller, Matthew R. Aronoff, Ronald T. Raines

Brett VanVeller

1-Dimethylamino-8-methylaminonaphthalene is put forth as a protecting group for benzoxaboroles. The ensuing complex is fluorescent, charge-neutral, highly stable under basic conditions, stable to anhydrous acid, and readily cleavable in aqueous acid to return the free benzoxaborole.

Stomatin-Like Protein 2 Deficiency In T Cells Is Associated With Altered Mitochondrial Respiration And Defective Cd4+ T Cell Responses., Darah A Christie, Panagiotis Mitsopoulos, Julianna Blagih, Stanley D Dunn, Julie St-Pierre, Russell G Jones, Grant M Hatch, Joaquín Madrenas

Stanley D Dunn

Stomatin-like protein 2 (SLP-2) is a mostly mitochondrial protein that regulates mitochondrial biogenesis and function and modulates T cell activation. To determine the mechanism of action of SLP-2, we generated T cell-specific SLP-2-deficient mice. These mice had normal numbers of thymocytes and T cells in the periphery. However, conventional SLP-2-deficient T cells had a posttranscriptional defect in IL-2 production in response to TCR ligation, and this translated into reduced CD4(+) T cell responses. SLP-2 deficiency was associated with impaired cardiolipin compartmentalization in mitochondrial membranes, decreased levels of the NADH dehydrogenase (ubiquinone) iron-sulfur protein 3, NADH dehydrogenase (ubiquinone) 1β subcomplex subunit …

Comparative Metabolism Of Phenanthro[3,4-B]Thiophene And Benzo[C]Phenanthrene, Jaquan M. Williams

Jaquan M Williams

ABSTRACT OF THESIS Comparative Metabolism of Phenanthro[3,4-b]Thiophene And Benzo[c]Phenanthrene Polycyclic aromatic hydrocarbons (PAHs) and their sulfur-heterocyclic analogs (thia-PAHs) are commonly occurring persistent environmental contaminants formed by incomplete combustion of organic matter. A number of thia-PAHs have shown significant mutagenic and carcinogenic activities. As noted with PAHs, these chemical contaminants also require metabolic activation in order to exhibit their mutagenic and carcinogenic effects. In the present study, a comparison of the metabolism of highly mutagenic phenanthro[3,4-b]thiophene (P[3,4-b]T) and its weakly mutagenic carbon analogue, benzo[c]phenanthrene (B[c]P), was investigated. Metabolism studies were conducted using liver microsomes from induced rats, un-induced rats, as well …

Longitudinal Transcriptomic Dysregulation In The Peripheral Blood Of Transgenic Huntington’S Disease Monkeys, Jannet Kocerha, Yuhong Liu, David Willoughby, Kumaravel Chidamparam, Joseph Benito, Kate Nelson, Yan Xu, Tim Chi, Heidi Engelhardt, Sean Moran, Shang-Hsun Yang, Shi-Hua Li, Xiao-Jiang Li, Katherine Larkin, Adam Neumann, Heather Banta, Jinjing Yang, Anthony W. S. Chan

Jannet Kocerha

Background: Huntington’s Disease (HD) is a progressive neurodegenerative disorder caused by an expansion in the polyglutamine (polyQ) region of the Huntingtin (HTT) gene. The clinical features of HD are characterized by cognitive, psychological, and motor deficits. Molecular instability, a core component in neurological disease progression, can be comprehensively evaluated through longitudinal transcriptomic profiling. Development of animal models amenable to longitudinal examination enables distinct disease-associated mechanisms to be identified.

Results: Here we report the first longitudinal study of transgenic monkeys with genomic integration of various lengths of the human HTT gene and a range of polyQ repeats. With this unique group …

Function And X-Ray Crystal Structure Of Escherichia Coli Yfde, Elwood A. Mullins, Kelly L. Sullivan, T. Joseph Kappock

T. Joseph Kappock

Many food plants accumulate oxalate, which humans absorb but do not metabolize, leading to the formation of urinary stones. The commensal bacterium Oxalobacter formigenes consumes oxalate by converting it to oxalyl-CoA, which is decarboxylated by oxalyl-CoA decarboxylase (OXC). OXC and the class III CoA-transferase formyl-CoA:oxalate CoA-transferase (FCOCT) are widespread among bacteria, including many that have no apparent ability to degrade or to resist external oxalate. The EvgA acid response regulator activates transcription of the Escherichia coli yfdXWUVE operon encoding YfdW (FCOCT), YfdU (OXC), and YfdE, a class III CoA-transferase that is ~30% identical to YfdW. YfdW and YfdU are necessary …

Cooperative Effects Of Drug-Resistance Mutations In The Flap Region Of Hiv-1 Protease, Jennifer Foulkes-Murzycki, Christina Rosi, Nese Yilmaz, Robert Shafer, Celia Schiffer

Celia A. Schiffer

Understanding the interdependence of multiple mutations in conferring drug resistance is crucial to the development of novel and robust inhibitors. As HIV-1 protease continues to adapt and evade inhibitors while still maintaining the ability to specifically recognize and efficiently cleave its substrates, the problem of drug resistance has become more complicated. Under the selective pressure of therapy, correlated mutations accumulate throughout the enzyme to compromise inhibitor binding, but characterizing their energetic interdependency is not straightforward. A particular drug resistant variant (L10I/G48V/I54V/V82A) displays extreme entropy-enthalpy compensation relative to wild-type enzyme but a similar variant (L10I/G48V/I54A/V82A) does not. Individual mutations of sites …

A High Omega-3 Fatty Acid Diet Has Different Effects On Early And Late Stage Myeloid Progenitors, Melinda Varney, James Buchanan, Yulia Dementieva, W. Hardman, Vincent Sollars

Yulia Dementieva

The effects of the polyunsaturated omega-3 (n-3) and omega-6 (n-6) fatty acids (FA) on hematopoiesis are complex in that both FA forms are processed into leukotrienes, eicosanoids, and prostaglandins, which can have independent effects. These FA have antagonistic effects in that n-6 FA prostaglandins tend to be pro-proliferative and pro-inflammatory, while the effects of n-3 FA prostaglandins are the opposite. We have previously shown that diets high in n-3 FA reduce the size of the middle to later stage myeloid progenitor compartment in FVB X sv129 F1hybrid mice. To assay the effects of high n-3 FA diets on earlier stages …

Global Analysis Of Gene Expression Changes During Retinoic Acid-Induced Growth Arrest And Differentiation Of Melanoma: Comparison To Differentially Expressed Genes In Melanocytes Vs Melanoma, Mary H. Estler, Goran Boskovic, James Denvir, Sarah Miles, Donald A. Primerano, Richard M. Niles

Goran Boskovic

BACKGROUND: The incidence of malignant melanoma has significantly increased over the last decade. Some of these malignancies are susceptible to the growth inhibitory and pro-differentiating effects of all-trans-retinoic acid (RA). The molecular changes responsible for the biological activity of RA in melanoma are not well understood. RESULTS: In an analysis of sequential global gene expression changes during a 4-48 h RA treatment of B16 mouse melanoma cells, we found that RA increased the expression of 757 genes and decreased the expression of 737 genes. We also compared the gene expression profile (no RA treatment) between non-malignant melan-a mouse melanocytes and …

N→Π* Interactions Of Amides And Thioamides: Implications For Protein Stability, Robert W. Newberry, Brett Vanveller, Ilia A. Guzei, Ronald T. Raines

Brett VanVeller

Carbonyl–carbonyl interactions between adjacent backbone amides have been implicated in the conformational stability of proteins. By combining experimental and computational approaches, we show that relevant amidic carbonyl groups associate through an n→π* donor–acceptor interaction with an energy of at least 0.27 kcal/mol. The n→π* interaction between two thioamides is 3-fold stronger than between two oxoamides due to increased overlap and reduced energy difference between the donor and acceptor orbitals. This result suggests that backbone thioamide incorporation could stabilize protein structures. Finally, we demonstrate that intimate carbonyl interactions are described more completely as donor–acceptor orbital interactions rather than dipole–dipole interactions.

Genetic Parameters For Concentrations Of Minerals In Longissimus Muscle And Their Associations With Palatability Traits In Angus Cattle, R. G. Mateescu, A. J. Garmyn, Richard G. Tait Jr., Qing Duan, Q. Liu, Mary S. Mayes, Dorian J. Garrick, A. L. Van Eenennaam, D. L. Vanoverbeke, G. G. Hilton, Donald C. Beitz, James M. Reecy

Richard G. Tait Jr.

The objective of this study was to estimate genetic parameters for concentrations of minerals in LM and to evaluate their associations with beef palatability traits. Samples of LM from 2,285 Angus cattle were obtained and fabricated into steaks for analysis of mineral concentrations and for trained sensory panel assessments. Nine minerals, including calcium, copper, iron, magnesium, manganese, phosphorus, potassium, sodium, and zinc, were quantified. Restricted maximum likelihood procedures were used to obtain estimates of variance and covariance components under a multiple-trait animal model. Estimates of heritability for mineral concentrations in LM varied from 0.01 to 0.54. Iron and sodium were …

Genetic Parameters For Carnitine, Creatine, Creatinine, Carnosine, And Anserine Concentration In Longissimus Muscle And Their Association With Palatability Traits In Angus Cattle, R. G. Mateescu, A. J. Garmyn, M. A. O'Neil, Richard G. Tait Jr., Almass A. Abuzaid, Mary S. Mayes, Dorian J. Garrick, A. L. Van Eenennaam, D. L. Vanoverbeke, G. G. Hilton, Donald C. Beitz, James M. Reecy

Richard G. Tait Jr.

The objective of this study was to estimate genetic parameters for carnitine, creatine, creatinine, carnosine, and anserine concentration in LM and to evaluate their associations with Warner-Bratzler shear force (WBSF) and beef palatability traits. Longissimus muscle samples from 2,285 Angus cattle were obtained and fabricated into steaks for analysis of carnitine, creatine, creatinine, carnosine, anserine, and other nutrients, and for trained sensory panel and WBSF assessments. Restricted maximum likelihood procedures were used to obtain estimates of variance and covariance components under a multiple-trait animal model. Estimates of heritability for carnitine, creatine, creatinine, carnosine, and anserine concentrations in LM from Angus …

Activities Of Methionine-Γ-Lyase In The Acidophilic Archaeon “Ferroplasma Acidarmanus” Strain Fer1, M. A. Khan, Madeline M. Lopez-Munoz, Charles W. Kaspar, Kai F. Hung

Kai F. Hung

Biogeochemical processes on exposed pyrite ores result in extremely high levels of sulfuric acid at these locations. Acidophiles that thrive in these conditions must overcome significant challenges, including an environment with proton concentrations at pH 3 or below. The role of sulfur metabolism in the archaeon “Ferroplasma acidarmanus” strain fer1’s ability to thrive in this environment was investigated due to its growth-dependent production of methanethiol, a volatile organic sulfur compound. Two putative sequences for methionine- γ-lyase (EC 4.4.1.11), an enzyme known to carry out α,γ-elimination on L-methionine to produce methanethiol, were identified in fer1. Bioinformatic analyses identified a conserved pyridoxal-5′-phosphate …

Tca Cycle Inactivation In Staphylococcus Aureus Alters Nitric Oxide Production In Raw 264.7 Cells, Chandirasegaran Massilamany, Arunakumar Gangaplara, Donald Gardner, James Musser, David Steffen, Greg Somerville, Jay Reddy

Greg A. Somerville

Inactivation of the Staphylococcus aureus tricarboxylic acid (TCA) cycle delays the resolution of cutaneous ulcers in a mouse soft tissue infection model. In this study, it was observed that cutaneous lesions in mice infected with wild-type or isogenic aconitase mutant S. aureus strains contained comparable inflammatory infiltrates, suggesting the delayed resolution was independent of the recruitment of immune cells. These observations led us to hypothesize that staphylococcal metabolism can modulate the host immune response. Using an in vitro model system involving RAW 264.7 cells, the authors observed that cells cultured with S. aureus aconitase mutant strains produced significantly lower amounts …

Acute Toxicity Of Copper Sulfate And Potassium Dichromate On Stygobiont Proasellus: General Aspects Of Groundwater Ecotoxicology And Future Perspectives, Ana Reboleira, Nelson Abrantes, Pedro Oromí, Fernando Gonçalves

Ana Sofia P.S. Reboleira

Karst systems harbor large groundwater resources for human consumption and represent an important habitat for rare and unprotected specialized animals, the so-called stygofauna. Due to the highly adapted features towards underground life, together with the geographic isolation provided by the subterranean aquifers, groundwater-dwelling animals may lose the ability to face sudden changes on their ecosystems, and therefore the risk of extinction is remarkably high. A little is known about their sensitiveness, especially linked to contamination pressure in urbanized karst areas. Understanding the impact of contaminants on stygofauna is important for setting groundwater environmental quality and management of karst systems. We …

Transmembrane Domain Three Contributes To The Ion Conductance Pathway Of Channelrhodopsin-2, Robert Dempski, Olga Gaiko

Robert E. Dempski

Channelrhodopsin-2 (ChR2) is a light-activated nonselective cation channel that is found in the eyespot of the unicellular green alga Chlamydomonas reinhardtii. Despite the wide employment of this protein to control the membrane potential of excitable membranes, the molecular determinants that define the unique ion conductance properties of this protein are not well understood. To elucidate the cation permeability pathway of ion conductance, we performed cysteine scanning mutagenesis of transmembrane domain three followed by labeling with methanethiosulfonate derivatives. An analysis of our experimental results as modeled onto the crystal structure of the C1C2 chimera demonstrate that the ion permeation pathway includes …

Psychosine, The Cytotoxic Sphingolipid That Accumulates In Globoid Cell Leukodystrophy, Alters Membrane Architecture, Jacqueline A. Hawkins-Salsbury, Archana R. Parameswar, Xuntian Jiang, Paul H. Schlesinger, Ernesto Bongarzone, Daniel S. Ory, Alexei V. Demchenko, Mark S. Sands

Alexei Demchenko

Inhibition Of Bacillus Cereus Growth By Bacteriocin Producing Bacillus Subtilis Isolated From Fermented Baobab Seeds (Maari) Is Substrate Dependent, Donatien Kaboré, Dennis S. Nielsen, Hagrétoui Sawadogo-Lingan, Bréhima Diawara, Mamoudou H. Dicko Prof., Mogens Jakobsen, Line Thorsen

Pr. Mamoudou H. DICKO, PhD

Quantitative Non-Isotopic Nitrocellulose Filter Binding Assays: Bacterial Mnsod-Dna Interactions, Joshua D. Czerwinski, David P. Mascotti

David P. Mascotti

No abstract provided.

Thermal And Sodium Dodecylsulfate Induced Transitions Of Streptavidin, Mark J. Waner, Irina Navrotskaya, Amanda Bain, Edward D. Oldham, David P. Mascotti

Mark J. Waner

The strong specific binding of streptavidin (SA) to biotin is utilized in numerous biotechnological applications. The SA tetramer is also known to exhibit significant stability, even in the presence of sodium dodecylsulfate (SDS). Despite its importance, relatively little is known about the nature of the thermal denaturation pathway for SA. This work uses a homogeneous SA preparation to expand on the data of previous literature reports, leading to the proposal of a model for temperature induced structural changes in SA. Temperature dependent data were obtained by SDS and native polyacrylamide gel electrophoresis (PAGE), differential scanning calorimetry (DSC), and fluorescence and …

The Effect Of Oxidant And The Non-Oxidant Alteration Of Cellular Thiol Concentration On The Formation Of Protein Mixed-Disulfides In Hek 293 Cells, Jasen Lee Gilge, Michael Fisher, Yuh-Cherng Chai

Yuh-Cherng Chai

Cellular molecules possess various mechanisms in responding to oxidant stress. In terms of protein responses, protein S-glutathionylation is a unique post-translational modification of protein reactive cysteines forming disulfides with glutathione molecules. This modification has been proposed to play roles in antioxidant, regulatory and signaling in cells under oxidant stress. Recently, the increased level of protein S-glutathionylation has been linked with the development of diseases. In this report, specific S-glutathionylated proteins were demonstrated in human embryonic kidney 293 cells treated with two different oxidative reagents: diamide and hydrogen peroxide. Diamide is a chemical oxidizing agent whereas hydrogen peroxide is a physiological …

Enhanced Transmission Of Higher Order Plasmon Modes With Random Au Nanoparticles In Periodic Hole Arrays, Fang-Tzu Chuang

Fang-Tzu Chuang

The enhanced transmission of higher order plasmon modes with random gold nanoparticles embedded in periodic hole arrays using asymmetric pair aperture as a unit is investigated in the midinfrared regime. Different thicknesses of gold film were deposited inside holes and then annealed to form randomly sized and distributed nanoparticles. The holes deposited with thin gold film exhibit significantly enhanced transmission in higher order modes after thermal annealing. The enormous local electric field around the nanoparticles enhances the scattering effect that contributes to the enhanced infrared transmission. This unique design, which integrates localized and propagating surface plasmons, provides an easy way …

Screening Of Commercial Cyclic Peptides As Inhibitor Envelope Protein Dengue Virus (Denv) Through Molecular Docking And Molecular Dynamics, Arli A. Parikesit, Kinanty Kinanty, Usman Sumo F. Tambunan

Arli A Parikesit

Dengue virus (DENV) has spread throughout the world, especially in tropical climates. Effective treatment of DENV infection is not yet available although several candidate vaccines have been developed. Treatment at this time is only to reduce symptoms and reduce the risk of death. Therefore, antiviral treatment is much needed. Envelope protein is one of the structural proteins of DENV which is known and could be a target of antiviral inhibitors and plays a special role in the fusion process. The aim of this research is to screen the commercial cyclic peptides which are used as inhibitors of envelope protein DENV …

Bone Morphogenetic Protein 4 Inhibition Of Tgfβ-2 Induced Cross-Linked Actin Networks In Human Trabecular Meshwork Cells, Rafael A. Lima, Michela Montecchi-Palmer, Jaclyn Bermudez, Abbot F. Clark Phd, Robert J. Wordinger Phd

ral5@rice.edu

Bone Morphogenetic Protein 4 Inhibition of TGFβ-2 Induced Cross-Linked Actin Networks (CLANs) in Human Trabecular Meshwork Cells

RAFAEL A. LIMA1, MICHELA MONTECCHI-PALMER2,3, JACLYN BERMUDEZ2 ABBOT F. CLARK2, AND ROBERT J. WORDINGER2

1 Rice University, Houston, Texas, 2 Department of Cell Biology and Anatomy, North Texas Eye Research Institute, University of North Texas Health Science Center, Fort Worth, Texas, 3 Alcon Research, Ltd., Fort Worth, Texas

Background:

Primary open-angle glaucoma (POAG) is a degenerative eye disease characterized by irreversible loss of retinal ganglion cells. One of the most significant risk factors for POAG is increased intraocular pressure (IOP). Elevated IOP is …

Advances In Profiling Of Noncoding Rnas In Neurological Disease, Murray J. Cairns, Jannet Kocerha

Jannet Kocerha

No abstract provided.

Transmembrane Serine Mutations Reduce The Minimum Pore Diameter Of Channelrhodopsin-2, Robert Dempski, Ryan Richards

Robert E. Dempski

Channelrhodopsin-2 (ChR2) is a microbial-type rhodopsin that, together with channelrhodopsin-1, mediates phototactic behavior in the green algae Chlamydomonas reinhardtii. Like all other microbial-type rhodopsins, ChR2 has seven transmembrane domains with the chromophore all-trans retinal bound to a single lysine residue. However, unlike other microbial-type rhodopsins, ChR2 functions as a non-selective cation channel and not an ion pump. A sequence alignment of ChR2 with the proton pump bacteriorhodopsin (bR) reveals that ChR2 lacks specific motifs within the transmembrane domains that facilitate non-covalent interactions and contribute to protein stability.

Periplasmic Response Upon Disruption Of Transmembrane Cu Transport In Pseudomonas Aeruginosa., José Argüello, Daniel Raimunda, Teresita Padilla-Benavides, Stefan Vogt, Sylvain Boutigny, Kaleigh Tomkinson, Lydia Finney

José M. Argüello

Pseudomonas aeruginosa, an opportunistic pathogen, has two transmembrane Cu(+) transport ATPases, CopA1 and CopA2. Both proteins export cytoplasmic Cu(+) into the periplasm and mutation of either gene leads to attenuation of virulence. CopA1 is required for maintaining cytoplasmic copper levels, while CopA2 provides copper for cytochrome c oxidase assembly. We hypothesized that transported Cu(+) ions would be directed to their destination via specific periplasmic partners and disruption of transport should affect the periplasmic copper homeostasis. Supporting this, mutation of either ATPase gene led to large increments in periplasmic cuproprotein levels. Toward identifying the proteins participating in this cellular response the …