Physical Sciences and Mathematics Commons^™

Can Acute Dermal Systemic Toxicity Tests Be Replaced With Oral Tests? A Comparison Of Route-Specific Systemic Toxicity And Hazard Classifications Under The Globally Harmonized System Of Classification And Labelling Of Chemicals (Ghs), Nigel P. Moore, David J. Andrew, Donald L. Bjerke, Stuart Creton, David Dreher, Thomas Holmes, Pilar Prieto, Troy Seidle, Tim G. Rowan

Troy Seidle, PhD

Acute systemic toxicity data (LD50 values) and hazard classifications derived in the rat following oral administration and dermal application have been analysed to examine whether or not orally-derived hazard classification or LD50 values can be used to determine dermal hazard classification. Comparing the oral and dermal classifications for 335 substances derived from oral and dermal LD50 values respectively revealed 17% concordance, and indicated that 7% of substances would be classified less severely while 76% would be classified more severely if oral classifications were applied directly to the dermal route. In contrast, applying the oral LD50 values within the dermal classification …

The Development Of New Concepts For Assessing Reproductive Toxicity Applicable To Large Scale Toxicological Programmes, S. Bremer, C. Pellizzer, S. Hoffmann, T. Seidle, T. Hartung

Troy Seidle, PhD

Large scale toxicological testing programmes which are currently ongoing such as the new European chemical legislation REACH require the development of new integrated testing strategies rather than applying traditional testing schemes to thousands of chemicals. The current practice of requiring in vivo testing for every possible adverse effect endanger the success of these programmes due (i) to limited testing facilities and sufficient capacity of scientific/technical knowledge for reproductive toxicity; (ii) an unacceptable number of laboratory animals involved (iii) an intolerable number of chemicals classified as false positive.

A key aspect of the implementation of new testing strategies is the determination …

Recent Advances In Quantum Chemical Predictions Of The Atmospheric Reactivity Of Criegee Intermediates, Keith Kuwata

Keith Kuwata

No abstract provided.

Inhibitors Of Nα-Acetyl-L-Ornithine Deacetylase: Synthesis, Characterization And Analysis Of Their Inhibitory Potency, J. Hlavacek, J. Picha, V. Vanek, J. Jiracek, J. Slaninova, V. Fucik, M. Budesinsky, Danuta Gillner, Richard Holz

Richard C. Holz

A series of N α-acyl (alkyl)- and N α-alkoxycarbonyl-derivatives of l- and d-ornithine were prepared, characterized, and analyzed for their potency toward the bacterial enzyme N α-acetyl-l-ornithine deacetylase (ArgE). ArgE catalyzes the conversion of N α-acetyl-l-ornithine to l-ornithine in the fifth step of the biosynthetic pathway for arginine, a necessary step for bacterial growth. Most of the compounds tested provided IC50 values in the μM range toward ArgE, indicating that they are moderately strong inhibitors. N α-chloroacetyl-l-ornithine (1g) was the best inhibitor tested toward ArgE providing an IC50 value of 85 μM while N α-trifluoroacetyl-l-ornithine (1f), N α-ethoxycarbonyl-l-ornithine (2b), and …

The Dape-Encoded N-Succinyl-L,L-Diaminopimelic Acid Desuccinylase From Haemophilus Influenzae Contains Two Active-Site Histidine Residues, Danuta M. Gillner, David L. Bienvenue, Boguslaw P. Nocek, Andzrej Joachimiak, Vincentos Zachary, Brian Bennett, Richard C. Holz

Richard C. Holz

The catalytic and structural properties of the H67A and H349A dapE-encoded N-succinyl-l,l-diaminopimelic acid desuccinylase (DapE) from Haemophilus influenzae were investigated. On the basis of sequence alignment with the carboxypeptidase from Pseudomonas sp. strain RS-16, both H67 and H349 were predicted to be Zn(II) ligands. The H67A DapE enzyme exhibited a decreased catalytic efficiency (180-fold) compared with wild-type (WT) DapE towards N-succinyldiaminopimelic acid. No catalytic activity was observed for H349A under the experimental conditions used. The electronic paramagnetic resonance (EPR) and electronic absorption data indicate that the Co(II) ion bound to H349A-DapE is analogous to that of WT DapE after the …

Mutation Of H63 And Its Catalytic Affect On The Methionine Aminopeptidase From Escherichia Coli, Sanghamitra Mitra, Brian Bennett, Richard C. Holz

Richard C. Holz

In order to gain insight into the mechanistic role of a flexible exterior loop near the active site, made up of Y62, H63, G64, and Y65, that has been proposed to play an important role in substrate binding and recognition in the methionyl aminopeptidase from Escherichia coli (EcMetAP-I), the H63A enzyme was prepared. Mutation of H63 to alanine does not affect the ability of the enzyme to bind divalent metal ions. The specific activity of H63A EcMetAP-I was determined using four different substrates of varying lengths, namely, l-Met-p-NA, MAS, MGMM and MSSHRWDW. For the smallest/shortest substrate (l-Met-p-NA) the specific activity …

Identification Of An Active Site-Bound Nitrile Hydratase Intermediate Through Single Turnover Stopped-Flow Spectroscopy, Natalie Gumataotao, Misty L. Kuhn, Natalia Hajnas, Richard C. Holz

Richard C. Holz

Stopped-flow kinetic data were obtained for the iron-type nitrile hydratase from Rhodococcus equi TG328-2 (ReNHase) using methacrylonitrile as the substrate. Multiple turnover experiments suggest a three-step kinetic model that allows for the reversible binding of substrate, the presence of an intermediate, and the formation of product. Microscopic rate constants determined from these data are in good agreement with steady state data confirming that the stopped-flow method used was appropriate for the reaction. Single turnover stopped-flow experiments were used to identify catalytic intermediates. These data were globally fit confirming a three-step kinetic model. Independent absorption spectra acquired between 0.005 …

Immobilization Of The Aminopeptidase From Aeromonas Proteolytica On Mg2+/Al3+ Layered Double Hydroxide Particles, Steven Frey, Stephanie Guilmet, Richard Egan, Alyssa Bennett, Sarah Soltau, Richard Holz

Richard C. Holz

A novel biomaterial formed by the immobilization of the Aminopeptidase from Aeromonas proteolytica (AAP) on synthetic Mg2+ and Al3+ ion-containing layered double hydroxide (LDH) particles was prepared. Immobilization of AAP on the LDH particles in a buffered, aqueous mixture is rapid such that the maximum loading capacity, 1 × 10−9 moles of AAP/mg LDH, is achieved in a few minutes. X-ray powder diffraction of LDH samples before and after treatment with AAP indicates that the enzyme does not intercalate between the layers of LDH, but instead binds to the surface. Treatment of AAP/LDH with various amounts of salt in a …

Analyzing The Binding Of Co(Ii)-Specific Inhibitors To The Methionyl Aminopeptidases From Escherichia Coli And Pyrococcus Furiosus, Sanghamitra Mitra, George Sheppard, Jieyi Wang, Brian Bennett, Richard Holz

Richard C. Holz

Methionine aminopeptidases (MetAPs) represent a unique class of protease that is capable of the hydrolytic removal of an N-terminal methionine residue from nascent polypeptide chains. MetAPs are physiologically important enzymes; hence, there is considerable interest in developing inhibitors that can be used as antiangiogenic and antimicrobial agents. A detailed kinetic and spectroscopic study has been performed to probe the binding of a triazole-based inhibitor and a bestatin-based inhibitor to both Mn(II)- and Co(II)-loaded type-I (Escherichia coli) and type-II (Pyrococcus furiosus) MetAPs. Both inhibitors were found to be moderate competitive inhibitors. The triazole-type inhibitor was found to …

Direct Patterning Of Silanized-Biomolecules On Semiconductor Surfaces, Dorjderem Nyamjav, Richard Holz

Richard C. Holz

A novel approach to pattern silanized-biomolecules directly onto glass (SiO_x) substrates via Dip-Pen nanolithography (DPN) and microcontact printing (μCP) is presented. Subsequent hybridization reactions of DPN patterned silanized-DNA with its complementary strands provide “proof-of-concept” that the patterned oligonucleotides maintain their biological activities. The fabrication strategy does not require premodification of substrates and offers a cheap and robust way to immobilize molecules on electronically important semiconductor surfaces.

Direct Patterning Of A Cyclotriveratrylene Derivative For Directed Self-Assembly Of C60, Zachary Osner, Dorjderem Nyamjav, Richard Holz, Daniel Becker

Richard C. Holz

A novel apex-modified cyclotriveratrylene (CTV) derivative with an attached thiolane-containing lipoic acid linker was directly patterned onto gold substrates via dip-pen nanolithography (DPN). The addition of a dithiolane-containing linker to the apex of CTV provides a molecule that can adhere to a gold surface with its bowl-shaped cavity directed away from the surface, thereby providing a surface-bound CTV host that can be used for the directed assembly of guest molecules. Subsequent exposure of these CTV microarrays to C60 in toluene resulted in the directed assembly of predesigned, spatially controlled, high-density microarrays of C60. The molecular recognition capabilities of this CTV …

Structural Basis For Catalysis By The Mono- And Dimetalated Forms Of The Dape-Encoded N-Succinyl-L,L-Diaminopimelic Acid Desuccinylase, Boguslaw Nocek, Danuta Gillner, Yao Fan, Richard Holz, Andzrej Joachimiak

Richard C. Holz

Biosynthesis of lysine and meso-diaminopimelic acid in bacteria provides essential components for protein synthesis and construction of the bacterial peptidoglycan cell wall. The dapE operon enzymes synthesize both meso-diaminopimelic acid and lysine and, therefore, represent potential targets for novel antibacterials. The dapE-encoded N-succinyl-l,l-diaminopimelic acid desuccinylase functions in a late step of the pathway and converts N-succinyl-l,l-diaminopimelic acid to l,l-diaminopimelic acid and succinate. Deletion of the dapE gene is lethal to Helicobacter pylori and Mycobacterium smegmatis, indicating that DapE's are essential for cell growth and proliferation. Since there are no similar pathways in humans, inhibitors …

Inhibitors Of Bacterial N-Succinyl-L,L-Diaminopimelic Acid Desuccinylase (Dape) And Demonstration Of In Vitro Antimicrobial Activity, Danuta Gillner, Nicola Armoush, Richard Holz, Daniel Becker

Richard C. Holz

The dapE-encoded N-succinyl-L,L-diaminopimelic acid desuccinylase (DapE) is a critical bacterial enzyme for the construction of the bacterial cell wall. A screen biased toward compounds containing zinc-binding groups (ZBG’s) including thiols, carboxylic acids, boronic acids, phosphonates and hydroxamates has delivered a number of micromolar inhibitors of DapE from Haemophilus influenzae, including the low micromolar inhibitor L-captopril (IC₅₀ = 3.3 μM, K_i = 1.8 μM). In vitro antimicrobial activity was demonstrated for l-captopril against Escherichia coli.

Lysine Biosynthesis In Bacteria: A Metallodesuccinylase As A Potential Antimicrobial Target, Danuta Gillner, Daniel P. Becker, Richard C. Holz

Richard C. Holz

In this review, we summarize the recent literature on dapE-encoded N-succinyl-l,l-diaminopimelic acid desuccinylase (DapE) enzymes, with an emphasis on structure–function studies that provide insight into the catalytic mechanism. Crystallographic data have also provided insight into residues that might be involved in substrate and hence inhibitor recognition and binding. These data have led to the design and synthesis of several new DapE inhibitors, which are described along with what is known about how inhibitors interact with the active site of DapE enzymes, including the efficacy of a moderately strong DapE inhibitor.

The Fe-Type Nitrile Hydratase From Comamonas Testosteroni Ni1 Does Not Require An Activator Accessory Protein For Expression In Escherichia Coli, Misty Kuhn, Salette Martinez, Natalie Gumataotao, Uwe Bornscheuer, Dali Liu, Richard Holz

Richard C. Holz

We report herein the functional expression of an Fe-type nitrile hydratase (NHase) without the co-expression of an activator protein or the Escherichia coli chaperone proteins GroES/EL. Soluble protein was obtained when the α- and β-subunit genes of the Fe-type NHase Comamonas testosteroni Ni1 (CtNHase) were synthesized with optimized E. coli codon usage and co-expressed. As a control, the Fe-type NHase from Rhodococcus equi TG328–2 (ReNHase) was expressed with (ReNHase+Act) and without (ReNHase−Act) its activator protein, establishing that expression of a fully functional, metallated ReNHase enzyme requires the co-expression of its activator protein, similar to all other Fe-type NHase enzymes reported …

Immobilization Of Motile Bacterial Cells Via Dip-Pen Nanolithography, Dorjderem Nyamjav, Sergey Rozhok, Richard Holz

Richard C. Holz

A strategy to bind bacterial cells to surfaces in a directed fashion via dip-pen nanolithography (DPN) is presented. Cellular attachment to pre-designed DPN generated microarrays was found to be dependent on the shape and size of the surface feature. While this observation is likely due in part to a dense, well formed mercaptohexadecanoic acid (MHA) monolayer generated via DPN, it may also simply be due to the physical shape of the surface structure. Motile Pseudomonas aeruginosa bacterial cells were observed to bind to DPN generated mercaptohexadecanoic acid/poly-L-lysine (MHA/PLL) line patterns, 'blocks' made up of eight lines with 100 nm spacings, …

Structural Characterization Of Zn(Ii)-, Co(Ii)-, And Mn(Ii)-Loaded Forms Of The Arge-Encoded N-Acetyl-L-Ornithine Deacetylase From Escherichia Coli, Ye Tao, Jacob Shokes, Wade Mcgregor, Robert Scott, Richard Holz

Richard C. Holz

The Zn, Co, and Mn K-edge extended X-ray absorption fine structure (EXAFS) spectra of the N-acetyl-l-ornithine deacetylase (ArgE) from Escherichia coli, loaded with one or two equivalents of divalent metal ions (i.e., [Zn(II)_(ArgE)], [Zn(II)Zn(II)(ArgE)], [Co(II)_(ArgE)], [Co(II)Co(II)(ArgE)], [Mn(II)_(ArgE)], and [Mn(II)Mn(II)(ArgE)]), were recorded. The Fourier transformed data (FT) for [Zn(II)_(ArgE)], [Zn(II)Zn(II)(ArgE)], [Co(II)_(ArgE)] and [Co(II)Co(II)(ArgE)] are dominated by a peak at 2.05 Å, that can be fit assuming five or six light atom (N,O) scatterers. Inclusion of multiple-scattering contributions from the outer-shell atoms of a histidine-imidazole ring resulted in reasonable Debye–Waller factors for these contributions and a slight reduction in the goodness-of-fit value …

Synthesis Of N-Succinyl-L,L-Diaminopimelic Acid Mimetics Via Selective Protection, V. Vanek, J. Picha, M. Budesinsky, M. Sanda, J. Jiracek, Richard Holz, J. Hlavacek

Richard C. Holz

The search for potential inhibitors that target so far unexplored bacterial enzyme mono-N-succinyl-L,Ldiaminopimelic acid desuccinylase (DapE) has stimulated a development of methodology for quick and efficient preparation of mono-N-acylated 2,6-diaminopimelic acid (DAP) derivatives bearing the different carboxyl groups or lipophilic moieties on their amino group.

Amphipathic Co-Oligomers For The Delivery Of Sirna, Christina Cooley, Erika Geihe, Robert Waymouth, Paul Wender

Christina B Cooley

Co-oligomer compounds, complexes of the same with polyanions, such as siRNAs, and methods for using the same are provided. the delivery of polynucleotides, into a cell. The subject co-oligomers include at least a liphopilic monomer and at least a hydrophilic monomer (e.g., a guanidinium containing monomer). In some embodiments, the co-oligomer compounds are capable of complexing a siRNA of interest, thereby increasing the cell permeability of the siRNA, prior to release of the siRNA into the cell. In some embodiments, the subject method is a method of delivery a siRNA into a cell. In some embodiments, the subject method is …

Conversion Of Red Fluorescent Protein Into A Bright Blue Probe, Oksana M. Subach, Illia S. Gundorov, Masami Yoshimura, Fedor V. Subach, Jinghang Zhang, David Grunwald, Ekaterina A. Souslova, Dmitriy M. Chudakov, Vladislav V. Verkhusha

David Grünwald

We used a red chromophore formation pathway, in which the anionic red chromophore is formed from the neutral blue intermediate, to suggest a rational design strategy to develop blue fluorescent proteins with a tyrosine-based chromophore. The strategy was applied to red fluorescent proteins of the different genetic backgrounds, such as TagRFP, mCherry, HcRed1, M355NA, and mKeima, which all were converted into blue probes. Further improvement of the blue variant of TagRFP by random mutagenesis resulted in an enhanced monomeric protein, mTagBFP, characterized by the substantially higher brightness, the faster chromophore maturation, and the higher pH stability than blue fluorescent proteins …

Composition And Optical Properties Of Secondary Organic Aerosol Particles, Felisha Imholt, Ryan Caylor, Matthew E. Wise, John Shilling

Matthew E. Wise

No abstract provided.

Optical Properties Of Secondary Organic Aerosols, Felisha Imholt, Ryan Caylor, Matthew E. Wise, John Shilling

Matthew E. Wise

Composition Of Secondary Organic Aerosols, Ryan Caylor, Felisha Imholt, Matthew E. Wise, John Shilling

Matthew E. Wise

A Generator-Produced Gallium-68 Radiopharmaceutical For Pet Imaging Of Myocardial Perfusion, Nigam Rath, Vijay Sharma, Jothilingam Sivapackiam, Scott E. Harpstrite, Julie L. Prior, Hannah Gu, David Piwnica-Worms

Nigam Rath

Magnetic Properties Of (Γ-Fe₂O₃)₈₀Ag₂₀ Nanocomposites Prepared In Reverse Micelles, Joan A. Wiemann, Jianbiao Dai, Jinke Tang, Gary J. Long, Leonard Spinu

Jinke Tang

The magnetic properties of nanoparticles of gamma-Fe2O3 prepared by reverse micelles have been studied by dc magnetization, transverse ac susceptibility, and Mössbauer spectroscopy. The nanoparticles of gamma-Fe2O3 in the nanocomposite (gamma-Fe2O3)80Ag20 exhibit superparamagnetic behavior. The blocking temperatures determined by the three methods indicate the superparamagnetic nature of (gamma-Fe2O3)80Ag20 above 70-80 K and show correlation with measuring time. The average particle diameter obtained by transmission electron microscopy of the gamma-Fe2O3 particles is ~10 nm and that of the Ag particles is ~20 nm. The average particle size determined from the magnetic analyses for the gamma-Fe2O3 particles is ~12 nm. Mössbauer spectra …

Magnetic Properties Of (Γ-Fe₂O₃)₈₀Ag₂₀ Nanocomposites Prepared In Reverse Micelles, Joan Wiemann, Jianbiao Dai, Jinke Tang, Gary Long, Leonard Spinu

Jinke Tang

The magnetic properties of nanoparticles of γ-Fe2O3 prepared by reverse micelles have been studied by dc magnetization, transverse ac susceptibility, and Mössbauer spectroscopy. The nanoparticles of γ-Fe2O3 in the nanocomposite (γ- Fe2 O3) 80Ag20 exhibit superparamagnetic behavior. The blocking temperatures determined by the three methods indicate the superparamagnetic nature of (γ- Fe2 O3) 80Ag20 above 70-80 K and show correlation with measuring time. The average particle diameter obtained by transmission electron microscopy of the γ-Fe2O3 particles is 10 nm and that of the Ag particles is 20 nm. The average particle size determined from the magnetic analyses for the γ-Fe2O3 …

The Analysis Of Muonium Hyperfine Interaction Measurements Of Thermal Rate Constants For Addition Reactions, Ronald Duchovic, A Wagner, R Turner, D Garner, D Fleming

Ronald J. Duchovic

A new model is developed for the analysis of muon spin relaxation (mu-SR) measurements of muonium (Mu) reaction rates with other chemical species. The kinetics component of the model is specifically developed for addition reactions but can be extended to direct reactions. The polarization component of the model assumes that the adduct formed possesses a single, effective spin precession frequency. The complete model is solved to obtain an analytic expression for the mu-SR signal as a function of time. Both a time-ordered sequence method and a Boltzmann spin equation method give equivalent solutions. These solutions are analyzed under various kinetic …

Understanding Life: Perspectives From Physical, Organic, And Biological Chemistry, Ronald Duchovic

Ronald J. Duchovic

No abstract provided.

Connecting Scholarship And Social Responsibility, Ronald Duchovic

Ronald J. Duchovic

No abstract provided.

The Potlib Library Of Potential Energy Functions And A New Global Analytic Pes For The H + O2 Reaction, Ronald Duchovic

Ronald J. Duchovic

No abstract provided.