Family Identification System for Electron Transfer Proteins Using a Motif Neural
Network Design. Pp. 9-16.
Cathy H. Wu, Sheng Zhao and Winona C. Barker
[Abstract]
The Binding of Cyclic Adenosine 3',5' Monophosphate to the Insulin Hexamer. Pp. 17-24.
Webe Kadima, Phila Raharivelomanana and Bethany Bechtel
[Abstract]
Controlling the Enantioselectivity of Sec-Alcohol Dehydrogenase from
Thermoanaerobacterium sp. KET4B1. Pp. 25-32.
Helen D. Simpson and Don A. Cowan
[Abstract]
Restriction of the Specificities of Trypsin and alpha-Chymotrypsin at a High Alkaline pH Value. Pp. 33-38.
Kenji Takahashi
[Abstract]
Purification of Recombinant Cholera Toxin Polypeptide A2 and Reconstitution with the Cholera Toxin B Subunit. Pp. 39-46.
Jameson A. McCann and William D. Picking
[Abstract]
High Resolution 1H NMR Spectra of Human Apolipoprotein C-III2 Using DPC to
Model the Lipoprotein Environment. Pp. 47-54.
Garry W. Buchko, Annett Rozek, Guangshun Wang, Jiri J. Frohlich and
Robert J. Cushley
[Abstract]
Evidence for the Identity of Rat Liver Microsomal Membrane-Bound Serine Proteinase with Hepsin. Pp. 55-62.
Hidemi Matsuzaki, Gwang-Ho Jeohn, Akihiro Iwamatsu, Yoshiaki Tamanoue, Yong-Tae Kim, Takayuki Takahashi and Kenji Takahashi
[Abstract]
Small Retro-Inverso Peptides Recognize MHC Class II HLA DR (alpha, beta1 *0401). Pp. 63-68.
Susan C. Howard, Michelle L. Zacheis, Christine P. Bono, Joseph K. Welply,
Gunnar J. Hanson, Jannifer L Vuletich, Lous J. Bedell, Neena L. Summers,
Benjamin D. Schwartz and Susan L. Woulfe
[Abstract]
Crystallisation Report
Purification, Crystallization and Preliminary X-Ray Analysis of a Phycourobilin-Containing Phycoerythrin. Pp. 69-74.
Stephan Ritter, Roger G. Hiller, Pamela M. Wrench, Thomas Wacker, Wolfram Welte and Kay Diederichs
[Abstract]
[Back to top] Development of a Capillary LC Blotting Systems for Low and Sub-Picomole Sequencing Sample Preparation. Kuo-Liang Hsi, Michael L. Kochersperger, William E. Werner, Chris H. Grimley, Lynn R. Zieske and Pau-Miau Yuan.
A capillary liquid chromatography (cLC)/microblotting system, 173A MicroBlotter, has recently been developed. This newly designed system consists of a capillary LC for sample separation and an on-line microblotter for direct collection of the separated peptides onto a strip of PVDF membrane. Applications using this new device for low and sub-picomole sample preparation and in conjunction with in-situ digestion techniques both in-gels or on-membrane will be demonstrated in this report.
[Back to top] Family Identification System for Electron Transfer Proteins Using a Motif Neural Network Design. Cathy H. Wu, Sheng Zhao and Winona C. Barker.
A neural network system, based on a motif identification neural design that incorporates both global and motif sequence information, has been developed to identify electron transfer protein families. The system was used to screen the SwissProt and PIR databases, and detected false negative family members missed by ProSite and PIR. The study shows the feasibility of developing a full system to assist database organization. The system is available from a WWW on-line server. New superfamily information will be incorporated into PIR.
[Back to top] The Binding of Cyclic Adenosine 3',5' Monophosphate to the Insulin Hexamer. Webe Kadima, Phila Raharivelomanana and Bethany Bechtel.
Evidence is presented for the binding of cyclic adenosine 3', 5' monophosphate (cAMP) to the insulin hexamer. Addition of cAMP to the T6 and R6 insulin hexamers, two allosteric forms of the hexamer, alters significantly the 1H NMR spectra of three aromatic residues that are located close to the surface of the insulin hexamer, indicating an interaction between the aromatic rings of cAMP and the protons of the insulin residues. The binding of cAMP to the insulin hexamer has never been documented, though cAMP is known to regulate secretion of insulin. This binding may have a role in the regulation of the function of the insulin hexamer in vivo, since the hexamer seem to have a specific binding site for cAMP.
[Back to top] Controlling the Enantioselectivity of Sec-Alcohol Dehydrogenase from Thermoanaerobacterium sp. KET4B1. Helen D. Simpson and Don A. Cowan.
The control of enantioselectivity by miscible organic solvents of a thermostable sec-ADH, purified from an anaerobic, thermophilic bacterium designated Thermoanaerobacterium sp. Ket4Bl, has been investigated. Experiments to compare the initial rates of oxidation of (R)-,and (S)-butan-2-ol in the presence of 0 to 40% (v/v) acetonitrile revealed that the VoR/VoS initial ratio increased with increasing solvent concentration. Other co-solvents were also found to influence the enantioselectivity but there was no direct correlation between either the log P or the dielectric constant of the solvent and the effect on the enantioselectivity.
[Back to top] Restriction of the Specificities of Trypsin and alpha-Chymotrypsin at a High Alkaline pH Value. Kenji Takahashi.
The specificities of trypsin and alpha-chymotrypsin at high alkaline pH values were investigated using dynorphin A and angiotensin I, respectively, as substrates. Under restricted conditions at pH 12.8, trypsin cleaved the Arg-X bonds (X =/= Pro) selectively without cleaving the Lys-X bonds, and a-chymotrypsin cleaved the Phe-X bond selectively without cleaving the Tyr-X bond. Therefore, trypsin and alpha-chymotrypsin may be generally useful for selective cleavages of Arg-X bonds and Phe-X bonds, respectively, at such a high pH value at which Lys and Tyr residues are unprotonated.
[Back to top] Purification of Recombinant Cholera Toxin Polypeptide A2 and Reconstitution with the Cholera Toxin B Subunit. Jameson A. McCann and William D. Picking.
Polypeptide A2 of cholera toxin (CTA2) was cloned, expressed and purified as a fusion protein possessing a leader with six tandem histidine residues. Following affinity purification, recombinant CTA2 was labeled with a fluorescent probe at its single cysteine and reconstituted with the cholera toxin B subunit (CTB) pentamer. This novel CTB/CTA2 complex will be useful for monitoring the position of the site of attachment of the cytotoxic Al polypeptide of cholera toxin relative to sites on CTB or GMl-containing membranes.
[Back to top] High Resolution 1H NMR Spectra of Human Apolipoprotein C-III2 Using DPC to Model the Lipoprotein Environment. Garry W. Buchko, Annett Rozek, Guangshun Wang, Jiri J. Frohlich and Robert J. Cushley.
While the apolipoproteins play a central biochemical role in lipid transportation and cardiovascular diseases, structural studies of the apolipoproteins have been primarily confined to optical spectroscopies (circular dichroism and fluorescence). Using deuterated dodecylphosphocholine (DPC) to model the lipoprotein environment, we report the acquisition of high resolution 1H NMR spectra for human apolipoprotein C-III2 isolated from the plasma of a type V hyperlipoproteinemic patient. At a peptide:DPC ratio of 1:60 the PKa of His18 was determined to be 6.15 suggesting the residue is solvent accessible.
[Back to top] Evidence for the Identity of Rat Liver Microsomal Membrane-Bound Serine Proteinase with Hepsin. Hidemi Matsuzaki, Gwang-Ho Jeohn, Akihiro Iwamatsu, Yoshiaki Tamanoue, Yong-Tae Kim, Takayuki Takahashi and Kenji Takahashi.
The serine proteinase previously isolated and partially characterized from the microsomal membrane fraction of rat liver was shown to be composed of two peptide chains of molecular masses of approximately 31 kDa and 19 kDa cross-linked by disulfide bond(s). Partial sequence analysis of peptides obtained by enzymatic hydrolysis indicated that the enzyme shares the same amino acid sequence with rat hepsin. These results provide evidence for the identity of these two enzymes.
[Back to top] Small Retro-Inverso Peptides Recognize MHC Class II HLA DR (alpha, beta1 *0401). Susan C. Howard, Michelle L. Zacheis, Christine P. Bono, Joseph K. Welply, Gunnar J. Hanson, Jannifer L Vuletich, Lous J. Bedell, Neena L. Summers, Benjamin D. Schwartz and Susan L. Woulfe.
We have synthesized a series of retro-inverso D-peptide analogues and a peptoid analog that mimic potent seven residue L-peptide ligands for DR(alpha, beta1*0401). The L-peptide ligands compete against binding of a 13 residue biotinylated ligand, HA307-319 (IC50 60nM), with competing peptide IC50s ranging from 30-200nM. The highest affinity heptamer retro-inverso D-peptide tested gave IC50 10 uM. No binding of the peptoid analog was detected.
[Back to top] Purification, Crystallization and Preliminary X-Ray Analysis of a Phycourobilin-Containing Phycoerythrin. Stephan Ritter, Roger G. Hiller, Pamela M. Wrench, Thomas Wacker, Wolfram Welte and Kay Diederichs.
Rhodophycean phycoerythrin (R-PE) was isolated from the red alga Grifflthsia monilis and purified on Sepharose. Untwinned crystals diffracting to 2.1A on a rotating anode X-ray source were obtained by the hanging-drop vapour diffusion method, and their space group determined as R3. SDS gel-electrophoresis of dissolved crystals showed the alpha and beta subunits of the protein (apparent molecular weight, about 19 kDa) and two different forms of the gamma-subunit (apparent molecular weight, 28 and 30 kDa) being present in the crystals.