... Mater. 2000, 12, 3642. [23] KL Jensen, AK Ganguly, J. Vac. Sci. Technol. ... Growth 2002, 237, 496. Enzyme Immobilization on Porous Silicon Surfaces** By Sonia E. LØtant, Bradley R. Hart, Staci R. Kane, Masood Z. Hadi, Sharon J.... more
... Mater. 2000, 12, 3642. [23] KL Jensen, AK Ganguly, J. Vac. Sci. Technol. ... Growth 2002, 237, 496. Enzyme Immobilization on Porous Silicon Surfaces** By Sonia E. LØtant, Bradley R. Hart, Staci R. Kane, Masood Z. Hadi, Sharon J. Shields, and John G. Reynolds* ...
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Photoluminescent (PL) porous silicon (PSi) has been modified for the attachment of biomolecules. Silicon wafers were etched by HF in CH 3 CH 2 OH and current to yield N-type PSi surfaces. A linker was constructed using hydrosilation... more
Photoluminescent (PL) porous silicon (PSi) has been modified for the attachment of biomolecules. Silicon wafers were etched by HF in CH 3 CH 2 OH and current to yield N-type PSi surfaces. A linker was constructed using hydrosilation reactions to give a direct Si-C bond on the surface. The other end of linker was designed so traditional protein cross-linking chemistry could be used to attach biomolecules. Dansyl cadaverine and Biotin-Streptavidin were attached to verify the utility of, and substantiate spectroscopically, the linking system. Glucuronidase was immobilized on the surface utilizing the linking system. The enzyme retained activity monitored through the conversion of p-nitro-phenyl-β-D-glucuronide top-nitro-phenol. The photoluminescence of the surface was retained, but varied upon enzymatic production of the p-nitro-phenol. Alteromonas sp. JD6.5 (OPAA-2) was also immobilized on the surface and exhibited sufficient enzymatic activity to transform p-nitro-phenyl-soman to p-nitro-phenol. The change in the photoluminescence of the surface was correlated to enzymatic activity producing the p-nitro-phenol hydrolysis product.
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ABSTRACT Chemical crosslinking is an important tool for probing protein structure and protein-protein interactions. The approach usually involves crosslinking of specific amino acids within a folded protein or protein complex, enzymatic... more
ABSTRACT Chemical crosslinking is an important tool for probing protein structure and protein-protein interactions. The approach usually involves crosslinking of specific amino acids within a folded protein or protein complex, enzymatic digestion of the crosslinked protein(s), and identification of the resulting crosslinked peptides by liquid chromatography/mass spectrometry (LC/MS). In this manner, distance constraints are obtained for residues that must be in close proximity to one another in the native structure or complex. As the complexity of the system under study increases, for example, a large multi-protein complex, simply measuring the mass of a crosslinked species will not always be sufficient to determine the identity of the crosslinked peptides. In such a case, tandem mass spectrometry (MS/MS) could provide the required information if the data can be properly interpreted. In MS/MS, a species of interest is isolated in the gas phase and allowed to undergo collision induced dissociation (CID). Because the gas-phase dissociation pathways of peptides have been well studied, methods are established for determining peptide sequence by MS/MS. However, although crosslinked peptides dissociate through some of the same pathways as isolated peptides, the additional dissociation pathways available to the former have not been studied in detail. Software such as MS2Assign has been written to assist in the interpretation of MS/MS from crosslinked peptide species, but it would be greatly enhanced by a more thorough understanding of how these species dissociate. We are thus systematically investigating the dissociation pathways open to crosslinked peptide species. A series of polyalanine and polyglycine model peptides have been synthesized containing one or two lysine residues to generate defined inter- and intra-molecular crosslinked species, respectively. Each peptide contains 11 total residues, and one arginine residue is present at the carboxy terminus to mimic species generated by tryptic digestion. The peptides have been allowed to react with a series of commonly used crosslinkers such as DSS, DSG, and DST. The tandem mass spectra acquired for these crosslinked species are being examined as a function of crosslinker identity, site(s) of crosslinking, and precursor charge state. Results from these model studies and observations from actual experimental systems are being incorporated into the MS2Assign software to enhance our ability to effectively use chemical crosslinking in protein complex determination.
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Research Interests: Computational Biology, Fluorescence Microscopy, Biology, Tobacco, Membrane Proteins, and 15 moreCell Biology, Polysaccharides, Medicine, Multidisciplinary, Secretory Pathway, Endoplasmic Reticulum, Arabidopsis, Glucans, PLoS one, Reproducibility of Results, Xylans, Membrane Protein, Golgi Apparatus, Xyloglucan, and Plant Leaves
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Sandia National Laboratories Laboratory Directed Research and Development program contract no. DE-AC04-94AL85000
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Research Interests: Bioinformatics, Genetics, Algorithms, Computational Biology, Biology, and 15 moreDNA repair, Medicine, Disease susceptibility, Environmental Biotechnology, Molecular Epidemiology, Identification, Humans, Expressed Sequence Tags, BER, Gene, Active site, DNA damage response, Amino Acid Profile, Amino Acid Substitution Rates, and Base Excision Repair
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Synthetic biological engineering can be utilized to aide the advancement of improved long-term space flight. The potential to use synthetic biology as a platform to biomanufacture desired equipment on demand using the three dimensional... more
Synthetic biological engineering can be utilized to aide the advancement of improved long-term space flight. The potential to use synthetic biology as a platform to biomanufacture desired equipment on demand using the three dimensional (3D) printer on the International Space Station (ISS) gives long-term NASA missions the flexibility to produce materials as needed on site. Polyhydroxybutyrates (PHBs) are biodegradable, have properties similar to plastics, and can be produced in Escherichia coli using genetic engineering. Using PHBs during space flight could assist mission success by providing a valuable source of biomaterials that can have many potential applications, particularly through 3D printing. It is well documented that during PHB production E. coli cells can become significantly elongated. The elongation of cells reduces the ability of the cells to divide and thus to produce PHB. I aim to better understand cell division during PHB production, through the design, building, a...
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The liver rapidly and preferentially takes up Cu when administered orally or intravenously. Both kinetic and thermodynamic contributions may be involved. Kinetic factors are expected to determine the relative rates of Cu-transport by... more
The liver rapidly and preferentially takes up Cu when administered orally or intravenously. Both kinetic and thermodynamic contributions may be involved. Kinetic factors are expected to determine the relative rates of Cu-transport by different cell-types. Thermodynamic factors may help determine the amounts of copper accumulated at steady-state. Kinetic studies with hepatocytes suggestgd that albumin-Cu was not directly available for Cu-uptake. Histidine increased Cu-uptake1 from media which also contained albumin.l Apparently, the His2Cu complex which forms when histidine is in excess delivers Cu to a transport protein, and Cu is transported as the free ion. Cu-transport by fibroblasts is reported here as an example of Cu-transport by an extrahepatic cell type which is relevant to Cu-metabolism and Menkes disease.
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Extended abstract of a paper presented at Microscopy and Microanalysis 2012 in Phoenix, Arizona, USA, July 29 – August 2, 2012.
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Project Goals: Determine reliable, documented, and open process models of biofuel processes that can be used to guide research, investment, and policy. With the aim of understanding the contribution of enzymes to the cost of... more
Project Goals: Determine reliable, documented, and open process models of biofuel processes that can be used to guide research, investment, and policy. With the aim of understanding the contribution of enzymes to the cost of lignocellulosic biofuels, we constructed a technoeconomic model for the production of fungal cellulases. We found that the cost of producing enzymes was much higher than that commonly assumed in the literature, e.g. the cost contribution of enzymes to ethanol produced by the conversion of corn stover was found to be $0.68/gal if the sugars in the biomass could be converted at maximum theoretical yields, and $1.47/gal if the yields were based on saccharification and fermentation yields that have been previously reported in the scientific literature. We performed a sensitivity analysis to study the effect of feedstock prices and fermentation times on the cost contribution of enzymes to ethanol price. 2
Aindrila Mukhopadhyay,1,6* Edward Baidoo,1,6 Kelly Bender5,6 (bender@micro.siu.edu), Peter Benke,1,6 Swapnil Chhabra,1,6 Elliot Drury,3,6 Masood Hadi,2,6 Zhili He,4,6 Jay Keasling1,6 (keasling@.berkeley.edu), Kimberly Keller,3,6 Eric... more
Aindrila Mukhopadhyay,1,6* Edward Baidoo,1,6 Kelly Bender5,6 (bender@micro.siu.edu), Peter Benke,1,6 Swapnil Chhabra,1,6 Elliot Drury,3,6 Masood Hadi,2,6 Zhili He,4,6 Jay Keasling1,6 (keasling@.berkeley.edu), Kimberly Keller,3,6 Eric Luning,1,6 Francesco Pingitore,1,6 Alyssa Redding,1,6 Jarrod Robertson,3,6 Rajat Sapra,2,6 Anup Singh2,6 (aksingh@sandia.gov), Judy Wall3,6 (wallj@ missouri.edu), Grant Zane,3,6 Aifen Zhou,4,6 and Jizhong Zhou4,6 (jzhou@rccc.ou.edu)
Author(s): Hadi, Masood Z.; Tran, Huu; Morrison, Stephanie; Chu, Hou Cheng; DeGiovanni, A.; Adams, Paul D.
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Background. The VIMSS Computational Core group is tasked with data management, statistical analysis, and comparative and evolutionary genomics for the larger VIMSS effort. In the early years of this project, we focused on genome sequence... more
Background. The VIMSS Computational Core group is tasked with data management, statistical analysis, and comparative and evolutionary genomics for the larger VIMSS effort. In the early years of this project, we focused on genome sequence analysis including development of an operon prediction algorithm which has been validated across a number of phylogenetically diverse species. Recently, the Computational Core group has expanded its efforts, integrating large amounts of functional genomic data from several species into its comparative genomic framework.
Background: Metagenomics approaches provide access to environmental genetic diversity for biotechnology applications, enabling the discovery of new enzymes and pathways for numerous catalytic processes. Discovery of new glycoside... more
Background: Metagenomics approaches provide access to environmental genetic diversity for biotechnology applications, enabling the discovery of new enzymes and pathways for numerous catalytic processes. Discovery of new glycoside hydrolases with improved biocatalytic properties for the efficient conversion of lignocellulosic material to biofuels is a critical challenge in the development of economically viable routes from biomass to fuels and chemicals. Results: Twenty-two putative ORFs (open reading frames) were identified from a switchgrass-adapted compost community based on sequence homology to related gene families. These ORFs were expressed in E. coli and assayed for predicted activities. Seven of the ORFs were demonstrated to encode active enzymes, encompassing five classes of hemicellulases. Four enzymes were over expressed in vivo, purified to homogeneity and subjected to detailed biochemical characterization. Their pH optima ranged between 5.5 7.5 and they exhibit moderate ...
Procedures were developed to determine if live, adult two-spotted spidermites (Tetranychus urticae Koch) could be surface disinfested before being introduced into in vitro cultures of torenia (Torenia fournieri L.). Three time periods (5,... more
Procedures were developed to determine if live, adult two-spotted spidermites (Tetranychus urticae Koch) could be surface disinfested before being introduced into in vitro cultures of torenia (Torenia fournieri L.). Three time periods (5, 10, and 15 minutes) and five levels of sodium hypochlorite (0.05% to 0.25%) were evaluated. Surface disinfestation was accomplished by agitating 2 × 3 cm pieces of infested bean leaves in sodium hypochlorite solutions and then drying in a mite drier apparatus. All sodium hypochlorite concentrations disinfested the mites completely, however high concentration levels were lethal to the mites. Exposure periods of 10 and 15 minutes also significantly increased mortality. For optimum disinfestation of two-spotted spidermites with minimum mortality, a concentration of 0.05% sodium hypochlorite and 0.05% Tween-20 for 5 minutes should be used.
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A laboratory exercise on direct and indirect organogenesis from leaf explants is presented for students of plant tissue culture or plant propagation. Torenia fournieri, the wishbone flower, is used for this laboratory exercise because the... more
A laboratory exercise on direct and indirect organogenesis from leaf explants is presented for students of plant tissue culture or plant propagation. Torenia fournieri, the wishbone flower, is used for this laboratory exercise because the in vitro production of adventitious shoots from Torenia is easy to control, seeds are easy to obtain, and plants are easy to grow. Direct shoot organogenesis results from leaf explants without an intervening callus phase, and indirect shoot organogenesis is possible after 4 to 6 weeks of callus production from leaf explants. The basal medium for all forms of organogenesis contains Murashige and Skoog (MS) salts and vitamins, 30 g sucrose/liter, and 7 g agar/liter at pH 5.7. To obtain direct shoot organogenesis, leaf explants should be placed on the MS basal medium with 1.1 μM (0.25 mg·liter-1) 6-benzylaminopurine (BAP) and 0.25 μM (0.05 mg·liter-1) indole-3-butyric acid (IBA). If leaf explants are placed on MS medium with 2.3 μM (0.5 mg·liter-1) 2,...
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Research Interests: Bioinformatics, Evolutionary Biology, Earth Sciences, Environmental Science, Biogeochemistry, and 15 moreEnvironmental Remediation, Comparative Genomics, Imaging, Bioremediation, Biology, Metabolomics, Life Sciences, Extremophiles, Metagenomics, Functional Genomics, Environmental Sciences, Models, Microarrays, Environmental genomics, and Field Studies
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Research Interests: Environmental Science, Materials Science, Mass Transfer, Renewable Energy, Synthetic Biology, and 15 moreBioenergy, Energy Conservation, Cellulosic Ethanol, Cellulose, Cellulase, Fossil Fuels, Biofuel, Greenhouse Gas Emissions, Energy Intensity, Enzyme, Greenhouse gases, Pulp and Paper Industry, Energy Crop, Deconstruction of Building Materials, and Lignocellulosic Biomass
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CiteSeerX - Document Details (Isaac Councill, Lee Giles): null.
Research Interests: Engineering, Mechanical Engineering, Electrical Discharge Machining, Response Surface Methodology, Machining, and 15 moreExperimental Design, Mathematical Sciences, Factorial Design, Regression Analysis, Advanced manufacturing technology, Orthogonal Array, Cross Section, Analysis of Variance, Electric Discharge Machining, Material Removal Rate, Process Parameters, Corrosion Resistance, Heat affected zone, Regression equation, and Roundness
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We have characterized a β-glucuronosyltransferase (AtGlcAT14A) from Arabidopsis thaliana that is involved in the biosynthesis of type II arabinogalactan (AG). This enzyme belongs to the Carbohydrate Active Enzyme database... more
We have characterized a β-glucuronosyltransferase (AtGlcAT14A) from Arabidopsis thaliana that is involved in the biosynthesis of type II arabinogalactan (AG). This enzyme belongs to the Carbohydrate Active Enzyme database glycosyltransferase family 14 (GT14). The protein was localized to the Golgi apparatus when transiently expressed in Nicotiana benthamiana. The soluble catalytic domain expressed in Pichia pastoris transferred glucuronic acid (GlcA) to β-1,6-galactooligosaccharides with degrees of polymerization (DP) ranging from 3-11, and to β-1,3-galactooligosaccharides of DP5 and 7, indicating that the enzyme is a glucuronosyltransferase that modifies both the β-1,6- and β-1,3-galactan present in type II AG. Two allelic T-DNA insertion mutant lines showed 20-35% enhanced cell elongation during seedling growth compared to wild-type. Analyses of AG isolated from the mutants revealed a reduction of GlcA substitution on Gal-β-1,6-Gal and β-1,3-Gal, indicating an in vivo role of AtGlcAT14A in synthesis of those structures in type II AG. Moreover, a relative increase in the levels of 3-, 6- and 3,6-linked galactose (Gal) and reduced levels of 3-, 2- and 2,5-linked arabinose (Ara) were seen, suggesting that the mutation in AtGlcAT14A results in a relative increase of the longer and branched β-1,3- and β-1,6-galactans. This increase of galactosylation in the mutants is most likely caused by increased availability of the O6 position of Gal, which is a shared acceptor site for AtGlcAT14A and galactosyltransferases in synthesis of type II AG, and thus addition of GlcA may terminate Gal chain extension. We discuss a role for the glucuronosyltransferase in the biosynthesis of type II AG, with a biological role during seedling growth.