Heat Shock Proteins in Ascaris suum

Heat Shock Proteins in Ascaris suum

Date: August 1995
Creator: Chao, Sheng-Hao
Description: Ascaris suum were exposed to a number of stressors, including heavy metals and both high (40°C) and low (18°C) temperatures. The 70kD and 90kD heat shock proteins (HSPs) in the different A. suum tissues were analyzed by Western blot and quantitated by Macintosh Image Program.
Contributing Partner: UNT Libraries
Mechanisms of rapid receptive field reorganization in rat spinal cord

Mechanisms of rapid receptive field reorganization in rat spinal cord

Date: August 2002
Creator: Vu, Hung
Description: Rapid receptive field (RF) reorganization of somatosensory neurons in the rat dorsal horn was examined using extracellular single unit recording. Subcutaneous injection of lidocaine into RFs of dorsal horn neurons results in expansion of their RFs within minutes. The expanded RFs appear adjacent to or/and proximal to original RFs. Out of 63 neurons tested, 36 (58%) show RF reorganization. The data suggest that dorsal horn of spinal cord is one of the initial sites for RF reorganization. The neural mechanisms of this effect are not well understood. We propose that changes in biophysical properties (membrane conductance, length constant) of the neurons resulting from lidocaine injection contribute to RF reorganization. Iontophoretic application of glutamate onto dorsal horn neurons that show lidocaine induced RF's expansion were used to test the model. Application of glutamate produced reduction of reorganized RFs in 9 of 20 (45%) tested cells. Application of NBQX produced no effect on either original or expanded RFs indicate that RF shrinkage effects of glutamate involve NMDA receptors. The results are consistent with the prediction of the proposed model. Subcutaneous injection of capsaicin into tactile RFs of low threshold mechanoreceptive dorsal horn neurons produced no effect on the RF sizes that are ...
Contributing Partner: UNT Libraries
Subcellular Localization of N-acylphosphatidyl-ethanolamine Synthase in Cotyledons of Cotton Seedlings

Subcellular Localization of N-acylphosphatidyl-ethanolamine Synthase in Cotyledons of Cotton Seedlings

Date: December 1995
Creator: Sriparameswaran, Anuja
Description: N-acylation of phosphatidylethanolamine (PE) with free fatty acids catalyzed by N-acyl phosphatidylethanolamine (NAPE) synthase was reported in cotyledons of 24-h-old cotton seedlings. Here I report subcellular localization of this enzyme. Differential centrifugation, sucrose density gradient fractionation,aqueous two-phase partitioning and electron microscopy techniques were utilized to elucidate subcellular site(s) of NAPE synthase. Marker enzymes were used to locate organelles in subcellular fractions. Differential centrifugation indicated that NAPE synthase is present in more than one organelle and it is a membrane bound enzyme. Sucrose density gradient fractionations indicated that NAPE synthase is present in membranes derived from endoplasmic reticulum (ER),Golgi and possibly plasma membrane (PM) but not mitochondria, glyoxysomes or plastids. Aqueous two-phase partitioning experiments with cotton and spinach tissues supported these results but Goigi appeared to be the major site of NAPE synthesis. Electron microscopy of subcellular fractions was used to examine isolated fractions to provide visual confirmation of our biochemical results. Collectively, these results indicate that NAPE is synthesized in plant ER, Golgi and possibly PM.
Contributing Partner: UNT Libraries
Noncovalent crosslinking of SH1 and SH2 to detect dynamic flexibility of the SH1 helix

Noncovalent crosslinking of SH1 and SH2 to detect dynamic flexibility of the SH1 helix

Date: August 2000
Creator: Park, Hyunguk
Description: In this experiment, fluorescent N- (1-pyrenyl) iodoacetamide modified the two reactive thiols, SH1 (Cys 707) and SH2 (Cys 697) on myosin to detect SH1-SH2 a -helix melting. The excimer forming property of pyrene is well suited to monitor the dynamics of the SH1 and SH2 helix melting, since the excimer should only form during the melted state. Decreased anisotropy of the excimer relative to the monomeric pyrene fluorescence is consistent with the disordering of the melted SH1-SH2 region in the atomic model. Furthermore, nucleotide analogs induced changes in the anisotropy of the excimer, suggesting that the nucleotide site modulates the flexibility of SH1-SH2 region.
Contributing Partner: UNT Libraries
Nucleotide Inhibition of Glyoxalase II

Nucleotide Inhibition of Glyoxalase II

Date: May 1999
Creator: Gillis, Glen S
Description: The glyoxalase system mediates the conversion of methylglyoxal, a toxic ketoaldehyde, to D-lactic acid. The system is composed of two enzymes, glyoxalase I (Glo-I) and glyoxalase II (Glo-II), and exhibits an absolute requirement for a catalytic quantity of glutathione (GSH). Glo-I catalyzes the isomerization of a hemithioacetal, formed non-enzymatically from methylglyoxal and GSH, to the corresponding a -D-hydroxyacid thioester, s-D-lactoylglutathione (SLG). Glo-II catalyzes the irreversible breakdown of SLG to D-lactate and GSH. We have observed that ATP or GTP significantly inhibits the Glo-II activity of tissue homogenates from various sources. We have developed a rapid, one step chromatography procedure to purify Glo-II such that the purified enzyme remains "sensitive" to inhibition by ATP or GTP (Glo-II-s). Studies indicate that inhibition of Glo-II-s by nucleotides is restricted to ATP, GTP, ADP, and GDP, with ATP appearing most effective. Kinetics studies have shown that ATP acts as a partial non-competitive inhibitor of Glo-II-s activity, and further suggest that two kinetically distinguishable forms of the enzyme exist. The sensitivity of pure Glo-II-s to nucleotide inhibition is slowly lost on storage even at -80° C. This loss is accelerated at higher temperatures or in the presence of ATP. Kinetics studies on the resultant "insensitive" ...
Contributing Partner: UNT Libraries
Conformational Studies of Myosin and Actin with Calibrated Resonance Energy Transfer

Conformational Studies of Myosin and Actin with Calibrated Resonance Energy Transfer

Access: Use of this item is restricted to the UNT Community.
Date: May 2000
Creator: Xu, Jin
Description: Resonance energy transfer was employed to study the conformational changes of actomyosin during ATP hydrolysis. To calibrate the technique, the parameters for resonance energy transfer were defined. With conformational searching algorithms to predict probe orientation, the distances measured by resonance energy transfer are highly consistent with the atomic models, which verified the accuracy and feasibility of resonance energy transfer for structural studies of proteins and oligonucleotides. To study intramyosin distances, resonance energy transfer probes were attached to skeletal myosin's nucleotide site, subfragment-2, and regulatory light chain to examine nucleotide analog-induced structural transitions. The distances between the three positions were measured in the presence of different nucleotide analogs. No distance change was considered to be statistically significant. The measured distance between the regulatory light chain and nucleotide site was consistent with either the atomic model of skeletal myosin subfragment-1 or an average of the three models claimed for different ATP hydrolysis states, which suggested that the neck region was flexible in solution. To examine the participation of actin in the powerstroke process, resonance energy transfer between different sites on actin and myosin was measured in the presence of nucleotide analogs. The efficiencies of energy transfer between myosin catalytic domain and actin ...
Contributing Partner: UNT Libraries
Subcloning and Nucleotide Sequence of Two Positive Acting Regulatory Genes, xy1R and xy1S, from the Pseudomonas putida HS1 TOL Plasmid PDK1

Subcloning and Nucleotide Sequence of Two Positive Acting Regulatory Genes, xy1R and xy1S, from the Pseudomonas putida HS1 TOL Plasmid PDK1

Date: May 1992
Creator: Chang, Teh-Tsai
Description: TOL plasmids of Pseudomonas putida encode enzymes for the degradation of toluene and related aromatics. These genes are organized into two operons regulated by the Xy1R and Xy1S transcriptional activators. Previous analysis of the TOL pDK1 catechol-2,3-dioxygenase gene (xy1E) and a comparison of this gene to xy1E from the related TOL plasmid pWW0, revealed the existance of a substantial level of sequence homology (82%).
Contributing Partner: UNT Libraries
Application of Synthetic Peptides as Substrates for Reversible Phosphorylation

Application of Synthetic Peptides as Substrates for Reversible Phosphorylation

Date: August 1992
Creator: Abukhalaf, Imad Kazem
Description: Two highly homologous synthetic peptides MLC(3-13) (K-R-A-K-A-K-T-TK-K-R-G) and MLC(5-13) (A-K-A-K-T-T-K-K-R-G) corresponding to the amino terminal amino acid sequence of smooth muscle myosin light chain were utilized as substrates for protein kinase C purified from murine lymphosarcoma tumors to determine the role of the primary amino acid sequence of protein kinase C substrates in defining the lipid (phosphatidyl serine and diacylglycerol) requirements for the activation of the enzyme. Removal of the basic residues lysine and arginine from the amino terminus of MLC(3-13) did not have a significant effect on the Ka value of diacylglycerol. The binding of effector to calcium-protein kinase C appears to be random since binding of one effector did not block the binding of the other.
Contributing Partner: UNT Libraries