Biochemistry, Biophysics, and Structural Biology Commons^™

5-Hydroxydecanoate Is Metabolised In Mitochondria And Creates A Rate-Limiting Bottleneck For Β-Oxidation Of Fatty Acids, Peter J. Hanley, Stefan Dröse, Ulrich Brandt, Rachel A. Lareau, Abir L. Banerjee, D. K. Srivastava, Leonard J. Banaszak, Joseph J. Barycki, Paul P. Van Veldhoven, Jürgen Daut

Department of Biochemistry: Faculty Publications

5-Hydroxydecanoate (5-HD) blocks pharmacological and ischaemic preconditioning, and

has been postulated to be a specific inhibitor of mitochondrial ATP-sensitive K+ (KATP)

channels. However, recent work has shown that 5-HD is activated to 5-hydroxydecanoyl-CoA

(5-HD-CoA), which is a substrate for the first step of β-oxidation. We have now

analysed the complete β-oxidation of 5-HD-CoA using specially synthesised (and purified)

substrates and enzymes, as well as isolated rat liver and heart mitochondria, and compared

it with the metabolism of the physiological substrate decanoyl-CoA. At the second step of

β-oxidation, catalysed by enoyl-CoA hydratase, enzyme kinetics were …

Mobilization Of Intracellular Copper Stores By The Ctr2 Vacuolar Copper Transporter, Erin M. Rees, Jaekwon Lee, Dennis J. Thiele

Department of Biochemistry: Faculty Publications

Copper plays an essential role in processes including signaling to the transcription and protein trafficking machinery, oxidative phosphorylation, iron mobilization, neuropeptide maturation, and normal development. Whereas much is known about intracellular mobilization of ions such as calcium, little information is available on how eukaryotic cells mobilize intracellular copper stores. We describe a mechanism by which the Saccharomyces cerevisiae Ctr2 protein provides bioavailable copper via mobilization of intracellular copper stores. Whereas Ctr2 exhibits structural similarity to the Ctr1 plasma membrane copper importer, microscopic and biochemical fractionation studies localize Ctr2 to the vacuole membrane. We demonstrate that Ctr2 mobilizes vacuolar copper stores …

Interactions Between Small Heat Shock Protein Subunits And Substrate In Small Heat Shock Protein-Substrate Complexes, Kenneth L. Friedrich,, Kim C. Giese, Nicole R. Buan, Elizabeth Vierling

Department of Biochemistry: Faculty Publications

Small heat shock proteins (sHSPs) are dynamic oligomeric proteins that bind unfolding proteins and protect them from irreversible aggregation. This binding results in the formation of sHSP-substrate complexes from which substrate can later be refolded. Interactions between sHSP and substrate in sHSP-substrate complexes and the mechanism by which substrate is transferred to ATP-dependent chaperones for refolding are poorly defined. We have established C-terminal affinity-tagged sHSPs from a eukaryote (pea HSP18.1) and a prokaryote (Synechocystis HSP16.6) as tools to investigate these issues. We demonstrate that sHSP subunit exchange for HSP18.1 and HSP16.6 is temperature-dependent and rapid at the optimal growth temperature …

The Effects Of Intense Submicrosecond Electrical Pulses On Cells, Jingdong Deng, Karl H. Schoenbach, E. Stephen Buescher, Pamela S. Hair, Paula M. Fox, Stephen J. Beebe

Bioelectrics Publications

A simple electrical model for living cells predicts an increasing probability for electric field interactions with intracellular substructures of both prokaryotic and eukaryotic cells when the electric pulse duration is reduced into the submicrosecond range. The validity of this hypothesis was verified experimentally by applying electrical pulses (durations 100 μs– 60 ns, electric field intensities 3–150 kV/cm) to Jurkat cells suspended in physiologic buffer containing propidium iodide. Effects on Jurkat cells were assessed by means of temporally resolved fluorescence and light microscopy. For the longest applied pulses, immediate uptake of propidium iodide occurred consistent with electroporation as the cause of …

Real-Time Study Of Multidrug Resistance Mechanism In Pseudomonas Aeruginosa Using Nanoparticle Optics And Single Live Cell Imaging, Sophia Vasou Kyriacou

Chemistry & Biochemistry Theses & Dissertations

This thesis centers on the study of the xenobiotic efflux system in Pseudomonas aeruginosa, which is a ubiquitous bacterium. It resists many structurally and functionally diverse substrates due to expression of Mex-extrusion pumps, including MexAB-OprM, MexCD-OprJ, MexEF-OprN and MexXY-OprM systems. Despite extensive research, the structure and mechanism of multidrug resistance is unclear (1-9). For example, (i) how do MexA, MexB and OprM proteins assemble to extrude antibiotics? (ii) What is the antibiotic susceptibility of MexA, MexB, and OprM proteins? (iii) How do substrates cross the outer membrane of P. aeruginosa? (iv) Where are antibiotics accumulated inside the cell? This thesis …

Transmembrane Movement Of Exogenous Long-Chain Fatty Acids: Proteins, Enzymes, And Vectorial Esterification, Paul N. Black, Concetta C. Dirusso

Department of Biochemistry: Faculty Publications

INTRODUCTION......................................................................................................................................................454

FATTY ACIDS AND BIOLOGICAL MEMBRANES.............................................................................................455

Fatty Acid Transport Defined ...............................................................................................................................456

MODEL SYSTEMS TO INVESTIGATE FATTY ACID TRANSPORT ..............................................................456

Genetic Foundations of Fatty Acid Transport....................................................................................................456

PROTEINS IMPLICATED IN FATTY ACID TRANSPORT ...............................................................................456

Fatty Acid Translocase..........................................................................................................................................457

Fatty Acid Binding Protein—Membrane Bound ................................................................................................457

Fatty Acid Transport Protein...............................................................................................................................458

The Long-Chain Fatty Acid Transport Protein FadL .......................................................................................458

Fatty Acyl-CoA Synthetase ...................................................................................................................................459

VECTORIAL ACYLATION: ONE MECHANISM OPERATIONAL IN FATTY ACID TRANSPORT...........460

THE BACTERIAL PARADIGM ...............................................................................................................................460

Energetics of Fatty Acid Transport in Gram-Negative Bacteria......................................................................460

The Fatty Acid Transporter FadL........................................................................................................................461

The Fatty Acyl-CoA Synthetase FadD..................................................................................................................461

Structural Considerations of FACS .....................................................................................................................462

FATTY ACID TRANSPORT AND ACTIVATION IN YEAST: …

The Many Faces Of Vitamin B12: Catalysis By Cobalamin-Dependent Enzymes, Ruma Banerjee, Stephen W. Ragsdale

Department of Biochemistry: Faculty Publications

Vitamin B₁₂ is a complex organometallic cofactor associated with three subfamilies of enzymes: the adenosylcobalamin-dependent isomerases, the methylcobalamin-dependent methyltransferases, and the dehalogenases. Different chemical aspects of the cofactor are exploited during catalysis by the isomerases and the methyltransferases. Thus, the cobalt-carbon bond ruptures homolytically in the isomerases, whereas it is cleaved heterolytically in the methyltransferases. The reaction mechanism of the dehalogenases, the most recently discovered class of B₁₂ enzymes, is poorly understood. Over the past decade our understanding of the reaction mechanisms of B₁₂ enzymes has been greatly enhanced by the availability of large amounts of enzyme …

Electrically Mediated Delivery Of Vector Plasmid Dna Elicits An Antitumor Effect, L. Heller, D. Coppola

Bioelectrics Publications

In vivo electroporation is an efficient means of increasing plasmid DNA delivery to normal tissues, such as skin and muscle, as well as directly to tumors. In the experiments described here, plasmid DNA was delivered by in vivo electroporation to B16 mouse melanomas using two very different pulsing protocols. Reporter expression increased 21- or 42-fold, respectively with electroporation over injection alone. The growth of experimental melanomas with an approximate diameter of 4 mm on the day of treatment was monitored after electroporation delivery of reporter plasmid DNA. Remarkably, short-term complete regressions using one of these pulsing protocols occurred in up …

The Atp:Co(I)Rrinoid Adenosyltransferase (Coba) Enzyme Of Salmonella Enterica Requires The 2'-Oh Group Of Atp For Function And Yields Inorganic Triphosphate As Its Reaction Byproduct, Maris V. Fonseca, Nicole R. Buan, Alexander R. Horswill, Ivan Rayment, Jorge C. Escalante-Semerena

Department of Biochemistry: Faculty Publications

The specificity of the ATP:corrinoid adenosyltransferase (CobA) enzyme of Salmonella enterica serovar Typhimurium LT2 for its nucleotide substrate was tested using ATP analogs and alternative nucleotide donors. The enzyme showed broad specificity for the nucleotide base and required the 2’-OH group of the ribosyl moiety of ATP for activity. ³¹P NMR spectroscopy was used to identify inorganic triphosphate (PPP_i) as the byproduct of the reaction catalyzed by the CobA enzyme. Cleavage of triphosphate into pyrophosphate and orthophosphate did not occur, indicating that triphosphate cleavage was not required for release of the adenosylcorrinoid product. Triphosphate was a strong …

Vectorial Acylation In Saccharomyces Cerevisiae, Zhiying Zou, Fumin Tong, Nils J. Færgeman, Claus Børsting, Paul N. Black, Concetta Dirusso

Department of Biochemistry: Faculty Publications

In Saccharomyces cerevisiae Fat1p and fatty acyl-CoA synthetase (FACS) are hypothesized to couple import and activation of exogenous fatty acids by a process called vectorial acylation. Molecular genetic and biochemical studies were used to define further the functional and physical interactions between these proteins. Multicopy extragenic suppressors were selected in strains carrying deletions in FAA1 and FAA4 or FAA1 and FAT1. Each strain is unable to grow under synthetic lethal conditions when exogenous long-chain fatty acids are required, and neither strain accumulates the fluorescent long-chain fatty acid C₁-BODIPY-C₁₂ indicating a fatty acid transport defect. By using these …

The Atp:Co(I)Rrinoid Adenosyltransferase (Coba) Enzyme Of Salmonella Enterica Requires The 2’-Oh Group Of Atp For Function And Yields Inorganic Triphosphate As Its Reaction Byproduct, Maris V. Fonseca, Nicole R. Baun, Alexander R. Horswill, Ivan Rayment, Jorge C. Escalante-Semerena

Department of Biochemistry: Faculty Publications

The specificity of the ATP:corrinoid adenosyltransferase

(CobA) enzyme of Salmonella enterica serovar

Typhimurium LT2 for its nucleotide substrate was

tested using ATP analogs and alternative nucleotide donors.

The enzyme showed broad specificity for the nucleotide

base and required the 2’-OH group of the ribosyl

moiety of ATP for activity. ³¹P NMR spectroscopy

was used to identify inorganic triphosphate (PPP_i) as

the byproduct of the reaction catalyzed by the CobA

enzyme. Cleavage of triphosphate into pyrophosphate

and orthophosphate did not occur, indicating that

triphosphate cleavage was not required for release of

the adenosylcorrinoid product. Triphosphate was …

Studies On The Formation Of Dna-Cationic Lipid Composite Films And Dna Hybridization In The Composites, Murali Sastry, Vidya Ramakrishnan, Mrunalini Pattarkine, Krishna N. Ganesh

Faculty Works

The formation of composite films of double-stranded DNA and cationic lipid molecules (octadecylamine, ODA) and the hybridization of complementary single-stranded DNA molecules in such composite films are demonstrated. The immobilization of DNA is accomplished by simple immersion of a thermally evaporated ODA film in the DNA solution at close to physiological pH. The entrapment of the DNA molecules in the cationic lipid film is dominated by attractive electrostatic interaction between the negatively charged phosphate backbone of the DNA molecules and the protonated amine molecules in the thermally evaporated film and has been quantified using quartz crystal microgravimetry (QCM). Fluorescence studies …

Cationic Surfactant Mediated Hybridization And Hydrophobization Of Dna Molecules At The Liquid/Liquid Interface And Their Phase Transfer, Murali Sastry, Ashavani Kumar, Mrunalini Pattarkine, Vidya Ramakrishnan, Krishna N. Ganesh

Faculty Works

Hybridization of complementary oligonucleotides mediated by a cationic surfactant at the water/hexane interface leads to hydrophobic, double-helical DNA which may be readily phase transferred to the organic phase and cast into thin films on solid substrates.

Immobolisation And Biochemical Characterisation Of A Selected Commercially Available Laccase, Chandra Sundaralingam

Student Works (2000-2009)

ln this study, laccase from Novo Nordisk was chosen for biochemical characterization and immobilization investigation. The specific aims of this work was to develop optimum conditions for immobilization of laccase by entrapment method using copper alginate and characterize the laccase biochemically. The pH optimum of the crude enzyme extract was 5.5 and was found to be stable over a wide range of pH values. The apparent molecular weight of this enzyme was around 79,000 Daltons. Three peaks of activity (isoenzymes) were obtained from isoelectric focusing of pH values 4.7, 5.2 and 6.0. The optimum pH of the immobilized laccase by …

Hybridization Of Dna By Sequential Immobilization Of Oligonucleotides At The Air-Water Interface, Murali Sastry, Vidya Ramakrishnan, Mrunalini Pattarkine, Anand Gole, K. N. Ganesh

Faculty Works

The hybridization of DNA by sequential electrostatic and hydrogen-bonding immobilization of single-stranded complementary oligonucleotides at the air-water interface with cationic Langmuir monolayers is demonstrated. The complexation of the single-stranded DNA molecules with octadecylamine (ODA) Langmuir monolayers was followed in time by monitoring the pressure-area isotherms. A large (and slow) expansion of the ODA monolayer was observed during each stage of complexation in the following sequence: primary single-stranded DNA followed by complementary single-stranded DNA followed by the intercalator, ethidium bromide. Langmuir-Blodgett (LB) films of the ODA-DNA complex were formed on different substrates and characterized using quartz-crystal microgravimetry (QCM), Fourier transform infrared …

Electrically Mediated Plasmid Dna Delivery To Hepatocellular Carcinomas In Vivo, L. Heller, M. J. Jaroszeski, D. Coppola, C. Pottinger, R. Gilbert, Richard Heller

Bioelectrics Publications

Gene therapy by direct delivery of plasmid DNA has several advantages over viral gene transfer, but plasmid delivery is less efficient. In vivo electroporation has been used to enhance delivery of chemotherapeutic agents to tumors in both animal and human studies. Recently, this delivery technique has been extended to large molecules such as plasmid DNA. Here, the successful delivery of plasmids encoding reporter genes to rat hepatocellular carcinomas by in vivo electroporation is demonstrated.

Multiple Factors Independently Regulate Hila And Invasion Gene Expression In Salmonella Enterica Serovar Typhimurium, Robin L. Lucas, C. Phoebe Lostroh, Concetta C. Dirusso, Michael P. Spector, Barry L. Wanner, Catherine A. Lee

Department of Biochemistry: Faculty Publications

HilA activates the expression of Salmonella enterica serovar Typhimurium invasion genes. To learn more

about regulation of hilA, we isolated Tn5 mutants exhibiting reduced hilA and/or invasion gene expression. In

addition to expected mutations, we identified Tn5 insertions in pstS, fadD, flhD, flhC, and fliA. Analysis of the

pstS mutant indicates that hilA and invasion genes are repressed by the response regulator PhoB in the absence

of the Pst high-affinity inorganic phosphate uptake system. This system is required for negative control of the

PhoR-PhoB two-component regulatory system, suggesting that hilA expression may …

Multiple Factors Independently Regulate Hila And Invasion Gene Expression In Salmonella Enterica Serovar Typhimurium, Robin L. Lucas, C. Phoebe Lostroh, Concetta C. Dirusso, Michael P. Spector, Barry L. Wanner, Catherine A. Lee

Department of Biochemistry: Faculty Publications

HilA activates the expression of Salmonella enterica serovar Typhimurium invasion genes. To learn more

about regulation of hilA, we isolated Tn5 mutants exhibiting reduced hilA and/or invasion gene expression. In

addition to expected mutations, we identified Tn5 insertions in pstS, fadD, flhD, flhC, and fliA. Analysis of the

pstS mutant indicates that hilA and invasion genes are repressed by the response regulator PhoB in the absence

of the Pst high-affinity inorganic phosphate uptake system. This system is required for negative control of the

PhoR-PhoB two-component regulatory system, suggesting that hilA expression may …

Two-Step Processing Of Human Frataxin By Mitochondrial Processing Peptidase, Patrizia Cavadini, Jiri Adamec, Franco Taroni, Oleksandr Gakh, Grazia Isaya

Department of Biochemistry: Faculty Publications

We showed previously that maturation of the human

frataxin precursor (p-fxn) involves two cleavages by the

mitochondrial processing peptidase (MPP). This observation

was not confirmed by another group, however,

who reported only one cleavage. Here, we demonstrate

conclusively that MPP cleaves p-fxn in two sequential

steps, yielding a 18,826-Da intermediate (i-fxn) and a

17,255-Da mature (m-fxn) form, the latter corresponding

to endogenous frataxin in human tissues. The two cleavages

occur between residues 41–42 and 55–56, and both

match the MPP consensus sequence RX ↓ (X/S). Recombinant

rat and yeast MPP catalyze the pài step 4 and 40 …

Urea Hydrolysis Using Immobilized Urease, Anita Arullan

Student Works (2000-2009)

An inexpensive carrier matrix. vermiculite was used for immobilizing urease. A cost effective and simple method of coupling was developed using vermiculite. Immobilization of urease on Amberlite MB-land Nylon-6 tube were also studied. Activities of free and immobilized urease were studied and compared: Among all methods used for immobilization of urease. the method of coupling the enzyme to vermiculite gave the highest retained activity of 82% (Method I) and 89% (Method 2) at pH 6.0 and 5.5 respectively. Urease immobilized by Method I retained 69% and 30% of its original activity and Method 2 (using glutaraldehyde as coupling agent) retained …

Yeast And Human Frataxin Are Processed To Mature Form In Two Sequential Steps By The Mitochondrial Processing Peptidase, Steven S. Branda, Patrizia Cavadini, Jiri Adamec, Frantisek Kalousek, Franco Taroni, Grazia Isaya

Department of Biochemistry: Faculty Publications

Frataxin is a nuclear-encoded mitochondrial protein which is deficient in Friedreich’s ataxia, a hereditary neurodegenerative disease. Yeast mutants lacking the yeast frataxin homologue (Yfh1p) show iron accumulation in mitochondria and increased sensitivity to oxidative stress, suggesting that frataxin plays a critical role in mitochondrial iron homeostasis and free radical toxicity. Both Yfh1p and frataxin are synthesized as larger precursor molecules that, upon import into mitochondria, are subject to two proteolytic cleavages, yielding an intermediate and a mature size form. A recent study found that recombinant rat mitochondrial processing peptidase (MPP) cleaves the mouse frataxin precursor to the intermediate but not …

Anion Induced Blue To Purple Transition In Bacteriorhodopsin, Mrunalini Pattarkine, Anil K. Singh

Faculty Works

Purple membrane (PM, λ" role="presentation">λmax" role="presentation">max 570 nm) of H. halobium on treatment with sulphuric acid changes its colour to blue (λ" role="presentation">λmax" role="presentation">max 608 nm). The purple chromophore can be regenerated from the blue chromophore by exogeneous addition of anions such as CI−" role="presentation">− and HPO42−" role="presentation">2−4. Chloride ion is found to be more effective than the dibasic phosphate ion in regenerating the purple chromophore. Nevertheless, one thing common to the anion regeneration is that both CI−" role="presentation">− and HPO42−" role="presentation">2−4 show marked pH effect. At pH 1.0 the efficiency of …

Role Of The Escherichia Coli Fadr Regulator In Stasis Survival And Growth Phase-Dependent Expression Of The Uspa, Fad, And Fab Genes, Anne Farewell, Alfredo A. Diez, Concetta C. Dirusso, Thomas Nyström

Department of Biochemistry: Faculty Publications

The increased expression of the uspA gene of Escherichia coli is an essential part of the cell’s response to growth arrest. We demonstrate that stationary-phase activation of the uspA promoter is in part dependent on growth phase-dependent inactivation or repression of the FadR regulator. Transcription of uspA is derepressed during exponential growth in fadR null mutants or by including the fatty acid oleate in the growth medium of FadR1 cells. The results of DNA footprinting analysis show that FadR binds downstream of the uspA promoter in the noncoding region. Thus, uspA is a member of the fadR regulon. All …

Pattern Of Disease After Murine Hepatitis Virus Strain 3 Infection Correlates With Macrophage Activation And Not Viral Replication, M. Pope, O. Rotstein, E. Cole, S. Sinclair, Rebecca D. Parr, B. Cruz, R. Fingerote, S. Chung, R. Gorczynski, L. Fung, J. Leibowitz, Y. S. Rao, G. Levy

Faculty Publications

Murine hepatitis virus strain (MHV-3) produces a strain-dependent pattern of disease which has been used as a model for fulminant viral hepatitis. This study was undertaken to examine whether there was a correlation between macrophage activation and susceptibility or resistance to MHV-3 infection. Peritoneal macrophages were isolated from resistant A/J and susceptible BALB/cJ mice and, following stimulation with MHV-3 or lipopolysaccharide (LPS), analyzed for transcription of mRNA and production of interleukin-1 (IL-1), tumor necrosis factor alpha (TNF-alpha), transforming growth factor beta (TGF-beta), mouse fibrinogen-like protein (musfiblp), tissue factor (TF), leukotriene B4, and prostaglandin E2 (PGE2). Macrophages from BALB/cJ mice produced …

In Vivo Regulation Of Wheat-Leaf Phosphoenolpyruvate Carboxylase By Reversible Phosphorylation, Stephen M. G. Duff, Raymond Chollet

Department of Biochemistry: Faculty Publications

Regulation of C₃ phosphoenolpyruvate carboxylase (PEPC) and its protein-serine/threonine kinase (PEPC-PK) was studied in wheat (Trificum aesfivum) leaves that were excised from low-N-grown seedlings and subsequently illuminated and/or supplied with 40 mM KNO₃. The apparent phosphorylation status of PEPC was assessed by its sensitivity to i-malate inhibition at suboptimal assay conditions, and the activity state of PEPC-PK was determined by the in vitro ³²P labeling of purified maize dephospho-PEPC by [y³²PIATP/ Mg. lllumination (±NO₃^-) for 1 h led to about a 4.5-fold increase in the 50% inhibition constant for i-malate, which …

Molecular Cloning And Rare Cleavage Mapping Of Human 2p, 6q, 8q, 12q, And 18q Telomeres, Roberto A. Macina, Ken Morii, Xue-Lan Hu, Dimitri G. Negorev, Chrysanthe Spais, Lisa A. Ruthig, Harold C. Riethman

School of Medical Diagnostics & Translational Sciences Publications

Large terminal fragments of human chromosomes 2p, 6q, 8q, 12q, and 18q were cloned using yeast artificial chromosomes (YACs). RecA-assisted restriction endonuclease (RARE) cleavage analysis of genomic DNA samples from 11 unrelated individuals using YAC-derived probes confirmed the telomeric localizations of the half-YACs studied. The cloned Fragments provide telomeric closure of maps for the respective chromosome arms and will supply the reagents needed for analyzing and sequencing these distal subtelomeric regions.

Role Of Heat Shock Protein 70 Kda Cognate In Limiting Thermal Inactivation And Refolding Of Heat-Denatured Nuclear Type I Topoisomerase, Kuo-Kuang Wen

Theses and Dissertations in Biomedical Sciences

Previous studies (Ciavarra et al., 1994) demonstrated that the constitutive 70 kDa heat shock protein (hsc70) protected purified topoisomerase I from thermal injury. In addition, hsc70 was capable of regenerating catalytic activity of heat-denatured topoisomerase I. A whole cell lysate was also active in this reaction assay. The present study demonstrates that heat-denatured topoisomerase I is reactivated by a cytosolic fraction and that this activity is dependent on the presence of cytosolic hsc70. The efficacy of hsc70-mediated refolding of heat-denatured topoisomerase I is greatly enhanced by a cytosolic cofactor(s). In all these refolding reactions, exogenous ATP is not required. Size …

Characteristics Of Modified Leghemoglobins Lsolated From Soybean (Glycine Max Merr.) Root Nodules, Hyung-Kyun Jun, Gautam Sarath, Jose F. Moran, Manuel Becana, Robert V. Klucas, Fred W. Wagner

Department of Biochemistry: Faculty Publications

Hemoprotein derivatives of an abundant soybean (Glycine max Merr.) root nodule leghemoglobin, Lba, were studied for their modified spectral characteristics and physical properties. Three modified hemoprotein derivatives of Lba (Lbam₁, Lbam₂, and Lbam₃) were purified by preparative isoelectric focusing. The ferric forms of these pigments were green and exhibited anomalous spedra in the visible region as compared to the Lba³⁺ forms. These modified pigments showed a hypochromic shift of 10 nm for the charge transfer absorption maximum; however, differences were not apparent in the Sdret region. Upon binding with nicotinate, the α …

Sequence Of A Psac Gene From The Cyanobacterium Synechococcus Sp. Pcc 6301, Patricia L. Herman, Kartika Adiwilaga, John H. Golbeck, Donald P. Weeks

Department of Biochemistry: Faculty Publications

The psaC gene encodes PsaC, the apoprotein for the terminal iron-sulfur clusters, F_A and F_B, in the PSI reaction center of cyanobacteria, algae, and higher plants. PsaC functions as a membrane-bound oxidoreductase, accepting electrons from the F_x iron-sulfur cluster located on the PsaA/PsaB heterodimer and donating them to the soluble electron transfer proteins Fd and flavodoxin (reviewed in Bryant, 1992). The objective of our work is to clarify the role of PsaC in linear and cyclic electron transfer. The experimental organism is Syneckococcus sp. PCC 6301, a unicellular, freshwater cyanobacterium that is used extensively for physiological, …

Exopeptidase Catalyzed Site-Specific Bonding Of Supports, Labels And Bioactive Agents To Proteins, Fred W. Wagner, Thomas R. Coolidge, Dwane E. Wylie, Sheldon M. Schuster, William Lewis, Jay Stout

Department of Biochemistry: Faculty Publications

The invention provides a means for attaching a label, support or bioactive agent to a protein with an exopeptidase at a site that is remote from the active site of the protein. More specifically the invention is directed to a method for the attachment of an amino acid, amine and alcohol nucleophile to the carboxyl terminus of a protein. In one embodiment, a labeled nucleophile is attached to a protein such as an antibody. In other embodiments, the invention is directed to a method for the attachment of a protein to an immobilization support and to a method for the …