Reexamination of the Ergothioneine Biosynthetic Methyltransferase EgtD from Mycobacterium tuberculosis as a Protein Kinase Substrate

Ergothioneine has emerged as a crucial cytoprotectant in the pathogenic lifestyle of Mycobacterium tuberculosis. Production of this antioxidant from primary metabolites may be regulated by phosphorylation of Thr213 in the active site of the methyltransferase EgtD. The structure of mycobacterial EgtD suggests that this post-translational modification would require a large-scale change in conformation to make the active-site residue accessible to a protein kinase. In this report, we show that, under in vitro conditions, EgtD is not a substrate of protein kinase PknD.     

Effects of introduced aspartic and glutamic acid residues on the P'1 substrate specificity, pH dependence and stability of carboxypeptidase Y

Carboxypeptidase Y is a serine carboxypeptidase isolated from Saccharomyces cerevisiae with a preference for C-terminal hydrophobic amino acid residues. In order to alter the inherent substrate specificity of CPD-Y into one for basic amino acid residues in P'1, we have introduced Asp and/or Glu residues at a number of selected positions within the S'1 binding site. The effects of these substitutions on the substrate specificity, pH dependence and protein stability have been evaluated. The results presented here demonstrate that it is possible to obtain significant changes in the substrate preference by introducing charged amino acids into the framework provided by an enzyme with a quite different specificity. The introduced acidic amino acid residues provide a marked pH dependence of the (kcat/Km)FA-A-R-OH/(kcat/Km)FA-A-L-OH ratio. The change in stability upon introduction of Asp/Glu residues can be correlated to the difference in the mean buried surface area between the substituted and the substituting amino acid. Thus, the effects of acidic amino acid residues on the protein stability depend upon whether the introduced amino acid protrudes from the solvent accessible surface as defined by the surrounding residues in the wild type enzyme or is submerged below.     

Thick-layer glycerin-containing bichromated gelatin for recording volume holograms

A thick-layer light-sensitive material consisting of glycerin-containing bichromated gelatin has been prepared and used to record real-time volume holograms with 0.44 μm helium-cadmium laser radiation. The thickness of the layers is 400–600 μm. The holographic sensitivity of the material is 1 J/cm². The highest diffraction efficiency of holograms recorded using a symmetric system with parallel beams is 32%. Pis’ma Zh. Tekh. Fiz. 23, 62–65 (April 12, 1997)     

Muscle transformation of the sartorius muscle in a canine model: clinical impact for electrodynamic graciloplasty as a "neosphincter"

PURPOSE: Transformation of fast-twitching skeletal muscles to slow-twitching, slowly fatigable muscles has become of clinical interest in the recent past. Transposition and transformation of the gracilis muscle to use it as a substitute for a resected or defected anal sphincter (graciloplasty) have been reported as achieving promising results in the treatment of fecal incontinence caused by sphincter defects or following abdominoperineal anorectal excision for cancer. METHOD: This experimental study used a canine model and the sartorius muscle to evaluate the functional efficiency of two different configurations of the muscle loop to compare the presently applied transformation program (8 weeks) with a shorter (5 weeks) protocol. In six beagle dogs, both sartorius muscles were wrapped around two stomas, either in an alpha fashion or in the so-called split-sling technique. Muscle transformation was achieved by controlled neuromuscular stimulation either during eight (Program A) or five weeks (Program B). After completion of the transformation period, the function of the muscle slings was evaluated by manometry, and histomorphologic evaluation of the sartorius muscles was performed. RESULTS: It was shown that muscle transformation led to a slowly fatigable muscle that made it possible to perform continuos (tetanic) contraction, regardless of the configuration or the duration of the transformation. Median pressures created by these muscles also did not differ significantly. In accordance with these functional findings, the histologic evaluation showed the typical, significant increase of Type I fibers in both muscle slings and following both transformation protocols. Although the decrease of fast-twitching Type II fibers was more pronounced following the conventional (8 weeks) program, this finding did not influence the functional results. CONCLUSIONS: Results of our experiment indicate the possibility for using a shorter transformation protocol for transformation of the gracilis muscle during graciloplasty in the clinical setting. Furthermore, the efficacy and safety of the modified (split-sling) wrap technique was demonstrated.     

One-dimensional diffuser based on a thick layer of dichromated gelatin

A new diffuser capable of scattering light in one direction (one-dimensional diffuser) has been developed and experimentally tested. The diffuser is formed in a thick layer of dichromated gelatin. The scattering indicatrices of the diffuser measured in two mutually perpendicular directions are presented. It is demonstrated that the indicatrix width in one direction is significantly greater than that in the perpendicular direc-tion. However, no zero-order diffraction has been observed. The proposed one-dimensional diffuser exhibits close scattering indicatrices for reading at a wavelength of 0.44 and 0.63 μm.     

Prostaglandin synthase-1 and prostaglandin synthase-2 are coupled to distinct phospholipases for the generation of prostaglandin D2 in activated mast cells

Aggregation of IgE cell surface receptors on MMC-34 cells, a murine mast cell line, induces the synthesis and secretion of prostaglandin D2 (PGD2). Synthesis and secretion of PGD2 in activated MMC-34 cells occurs in two stages, an early phase that is complete within 30 min after activation and a late phase that reaches a maximum about 6 h after activation. The early and late phases of PGD2 generation are mediated by prostaglandin synthase 1 (PGS1) and prostaglandin synthase 2 (PGS2), respectively. Arachidonic acid, the substrate for both PGS1 and PGS2, is released from membrane phospholipids by the activation of phospholipases. We now demonstrate that in activated mast cells (i) secretory phospholipase A2 (PLA2) mediates the release of arachidonic acid for early, PGS1-dependent synthesis of PGD2; (ii) secretory PLA2 does not play a role in the late, PGS2-dependent synthesis of PGD2; (iii) cytoplasmic PLA2 mediates the release of arachidonic acid for late, PGS2-dependent synthesis of PGD2; and (iv) a cytoplasmic PLA2-dependent step precedes secretory PLA2 activation and is necessary for optimal PGD2 production by the secretory PLA2/PGS1-dependent early pathway.     

Berechnung der Wirbelstromdichten und-kräfte in der Umgebung großer supraleitender Spulen für den magnetischen Einschluß bei der Kernfusion

Übersicht Um die Betriebssicherheit großer supraleitender Magnetsysteme für die Kernfusionsforschung zu gewährleisten, muß die im Magnetsystem gespeicherte Energie schnell entladen werden können. Durch die veränderlichen großvolumigen Magnetfelder werden in benachbarten leitfähigen Strukturen durch Induktion hohe Wirbelströme hervorgerufen. Durch sie treten dort elektromagnetische Kräfte. auf, die einzelne Komponenten mechanisch überlasten können. Zur Berechnung dieser Kräfte wurde eine einfache numerischanalytische Methode entwickelt, die im folgenden vorgestellt und mit Finite-Elemente-Verfahren verglichen wird.

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Calculation of eddy current densities and forces around large superconducting coils for nuclear fusion

Contents The reliable operation of the superconducting magnet systems used for the magnetic confinement in nuclear fusion research requires the ability of a fast discharge of the stored magnetic energy. The corresponding magnetic field variation gives rise to high eddy currents by inductive coupling with surrounding conductive components. The eddy current forces can lead to a mechanical overload of some of these components. A simple numerical-analytical method was developed in order to calculate these forces. This method is presented and compared with the Finite Element Method.

     

Properties of permease dimer, a fusion protein containing two lactose permease molecules from Escherichia coli

An engineered fusion protein containing two tandem lactose permease molecules (permease dimer) exhibits high transport activity and is used to test the phenomenon of negative dominance. Introduction of the mutation Glu-325-->Cys into either the first or the second half of the dimer results in a 50% decrease in activity, whereas introduction of the mutation into both halves of the dimer abolishes transport. Lactose transport by permease dimer is completely inactivated by N-ethylmaleimide; however, 40-45% activity is retained after N-ethylmaleimide treatment when either the first or the second half of the dimer is replaced with a mutant devoid of cysteine residues. The observations demonstrate that both halves of the fusion protein are equally active and suggest that each half may function independently. To test the possibility that oligomerization between dimers might account for the findings, a permease dimer was constructed that contains two different deletion mutants that complement functionally when expressed as untethered molecules. Because this construct does not catalyze lactose transport to any extent whatsoever, it is unlikely that the two halves of the dimer interact or that there is an oligomeric interaction between dimers. The approach is consistent with the contention that the functional unit of lactose permease is a monomer.     

Der PC in zehn Jahren und seine Auswirkungen auf die Gesellschaft

In this article it is argued that in ten years’ time personal computers as we know them now won’t exist anymore — all their functions will by then be integrated in mobiles which will be our steady companions. As such, mobiles will change our lives to an enormous extent. Technology as well as consequences will be discussed in this paper.