Molecular cloning and expression of human trophoblast antigen FDO161G and its identification as 3 beta-hydroxy-5-ene steroid dehydrogenase

The monoclonal antibody FDO161G reacts with a 43-kDa protein found in human extravillous trophoblast, syncytiotrophoblast, adrenal cortex, interstitial cells of the testis and ovarian follicle cumulus cells. cDNAs for this protein have been isolated from the lambda gt11 library, sequenced, and expressed in COS-7 cells. The protein was identified as 3 beta-hydroxy-5-ene steroid dehydrogenase (HSD). The sequence of the HSD protein raises questions about its association with cell membrane systems. The lack of reactivity of FDO161G with other tissues suggests that HSD has a limited tissue distribution and that other enzymes may exist in peripheral tissues, which can convert delta 5 3-hydroxysteroids to delta 4 3-ketosteroids.     

Atypical nucleosome spacing of rat neuronal identifier elements in non-neuronal chromatin.

Rat neuronal identifier (ID) elements are located in chromatin regions that are organized in nucleosomal structures in both neuronal and non-neuronal cells. A subpopulation of ID sequences in chromatin of liver and kidney cells are relatively resistant to micrococcal nuclease digestion and are organized in nucleosomes exhibiting an atypically short repeat length. Other repetitive elements do not show this organization.     

Selective solubilization of xylan in pulp using a purified xylanase fromTrichoderma harzianum

Summary The specificity of action of a cellulase-free xylanase preparation on pulp fibers was revealed by the composition of the solubilized products after enzyme treatment. The neutral carbohydrates released by the treatment of two hardwood kraft pulps were composed exclusively of xylooligomers. A similar treatment of Solka Floc showed no detrimental effect on the degree of polymerization of the cellulose fibers, as determined by size exclusion chromatographic analysis.     

T cell responses to cleaved rabies virus glycoprotein and to synthetic peptides

The antigenic structure of the rabies virus glycoprotein has been studied. A limited number of fragments were obtained by cyanogen bromide (CNBr) cleavage of viral glycoprotein, and eight large peptides were isolated by using sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. These were tested for their capacity to stimulate the proliferation of nylon wool-purified T cells obtained from spleens of rabies-immune A/J mice. Three peptides (Cr1, Cr2 plus Cr2A, and Cr3) stimulated antigen-specific proliferation, indicating that at least three T cell determinants of the native molecule are sequential or continuous in nature. Stimulation was also obtained with 27-residue and 13-residue synthetic peptides (designated R21 and R20, respectively) that included sequences towards the carboxy terminal end of Cr1, but not with synthetic peptides that included sequences of Cr2 and Cr3 (which are both glycosylated in virus-derived material). The intact viral glycoprotein and synthetic peptide R21 stimulated T lymphocytes with surface characteristics of helper cells, and induced the production of interleukin 2 by these lymphocytes. Synthetic peptides R20 and R21 also stimulated a minor population of Lyt-2-positive cells, which were not yet identified as either suppressor or cytotoxic T lymphocytes.     

Solution structure of neuronal bungarotoxin determined by two-dimensional NMR spectroscopy: calculation of tertiary structure using systematic homologous model building, dynamical simulated annealing, and restrained molecular dynamics.

Neuronal bungarotoxin has previously been shown, using two-dimensional 1H NMR spectroscopy, to have a triple-stranded antiparallel beta-sheet structure which dimerizes in solution [Oswald, R.E., Sutcliffe, M.J., Bamberger, M., Loring, R.H., Braswell, E., & Dobson, C.M. (1991) Biochemistry 30, 4901-4909]. In this paper, structural calculations are described which use the 582 experimentally measured NOE restraints in conjunction with 27 phi-angle restraints from J-value measurements. The positions of the N-terminal region and C-terminal region were poorly defined in the calculated structures with respect to the remainder of the structure. The region of the structure containing the triple-stranded beta-sheet was, however, well defined and similar to that found in the structure of homologous alpha-bungarotoxin (45% amino acid identity). The experimental restraints did not result in a well-defined dimer interface region because of the small number of NOEs which could be identified in this region. An approach was therefore adopted which produced model structures based to varying degrees on the alpha-bungarotoxin structure. Fourteen different structures were generated in this manner and subsequently used as starting points for refinement using dynamical simulated annealing followed by restrained molecular dynamics. This approach, which combines NMR data and homologous model building, has enabled a family of structures to be proposed for the dimeric molecule. In particular, Phe49 has been identified as possibly playing an important role in dimer formation, this residue in one chain interacting with the corresponding residue in the adjacent chain.     

Organization and nucleotide sequence of the Streptococcus mutans galactose operon

The galactose operon encoding a repressor and genes for the Leloir pathway for galactose metabolism (galactokinase, galactose-1-phosphate-uridyl transferase and UDP glucose-4-epimerase) was located adjacent to the multiple sugar metabolism (msm) operon on the chromosome of Streptococcus mutans Ingbritt (serotype c) and the complete nucleotide sequence of this 5-kilobase region was determined. The Leloir pathway was induced by the presence of galactose in the growth medium or following the release of intracellular galactose after uptake and cleavage of -galactosides by the multiple sugar metabolism system. Analysis of the mechanism of galactose transport confirmed the absence of a galactose-specific phosphotransferase system and suggested the presence of an inducible galactose permease. Evidence is presented that galactose transport is independent of the proton motive force and may be ATP-dependent.     

Three-dimensional models of non-NMDA glutamate receptors.

Structural models have been produced for three types of non-NMDA inotropic glutamate receptors: an AMPA receptor, GluR1, a kainate receptor, GluR6; and a low-molecular-weight kainate receptor from goldfish, GFKAR alpha. Modeling was restricted to the domains of the proteins that bind the neurotransmitter glutamate and that form the ion channel. Model building combined homology modeling, distance geometry, molecular mechanics, interactive modeling, and known constraints. The models indicate new potential interactions in the extracellular domain between protein and agonists, and suggest that the transition from the "closed" to the "open" state involves the movement of a conserved positive residue away from, and two conserved negative residues into, the extracellular entrance to the pore upon binding. As a first approximation, the ion channel domain was modeled with a structure comprising a central antiparallel beta-barrel that partially crosses the membrane, and against which alpha-helices from each subunit are packed; a third alpha-helix packs against these two helices in each subunit. Much, but not all, of the available data were consistent with this structure. Modifying the beta-barrel to a loop-like topology produced a model consistent with available data.     

MsmE, a lipoprotein involved in sugar transport in Streptococcus mutans.

Metabolic labelling by [14C]palmitic acid showed that growth of Streptococcus mutans LT11 in raffinose, an inducer of the msm operon, resulted in increased production of a 45-kDa lipoprotein corresponding to MsmE, which is believed to be a sugar-binding protein. MsmE was also labelled when an msmE clone was expressed in Escherichia coli. The presence of a lipid anchor on MsmE provides a likely explanation of how the sugar-binding protein component of the msm binding protein-dependent multiple sugar transport system is retained at the cell surface.