Proton NMR resonance assignments and surface accessibility of tryptophan residues of a dimeric phospholipase A2 from Trimeresurus flavoviridis

Proton NMR spectra of a dimeric phospholipase A₂ from Trimeresurus flavoviridis have been recorded. N-1 proton resonances of the tryptophan indole rings have been detected and assigned to specific positions, Trp-3/Trp-30, Trp-68 and Trp-108, by comparing the spectra of the enzyme derivatives with tryptophans oxidized to differing extents. Photo-CIDNP experiments have revealed that Trp-68 and Trp-108 are exposed while Trp-3 and Trp-30 are buried in the molecule. This is consistent with the X-ray crystal structure of a homologous phospholipase A₂ from Crotalus atrox where residues 3 and 30 are located at a dimer interface, but inconsistent with the results of stepwise oxidation of tryptophan residues.     

On-off regulation of gene expression from tryptophan promoter with cross-flow filtration

Summary Recombinant plasmid containing β-galactosidase gene fused to trp promoter (pMCT98) and that containing cloned trp repressor gene (pRLK13) were introduced into Escherichia coli C600. The bacterium was cultivated in a jar-fermetor equipped with a cross-flow filtration apparatus to attain the on-off regulation of the gene expression by controlling tryptophan concentration in the medium. In logarithmic growth phase, the cross-flow filtration was started. Tryptophan concentration dropped to a low level within 1 h and an efficient expression of β-galactosidase gene was started. By this twostage cultivation, very high biomass was achieved (final OD₅₇₀: 150) and the amount of produced β-galactosidase was about 10% of total cellular proteins.     

Some properties of randomly connected networks of neuron-like elements with refractory

Dynamic properties of randomly connected networks consisting of neuron-like elements with refractory are investigated from a macroscopic point of view. Equations describing the transition of the activity level of the network — a macroscopic state — are derived under some hypotheses on the stochastic properties of the network. The equations are characterized by a set of parameters which are determined by distributions of the threshold values of elements and the weighting values of connection between elements. It is shown that a network behaves like a monostable, bistable or astable circuit when its refractory period is less than one time unit and that a network is monostable or bistable when its refractory period is longer than two time units. An oscillatory network, on the other hand, is always realized if the network has a feedback mechanism which decreases the excitability of neurons when high activity level is sustained. Some results of computer simulation of randomly connected neuron networks are also presented.     

Clinical Studies with Mazindol

An anoerxiant, mazindol suppresses food intake by 1)stimulating β-adrenergic receptors, 2)inhibiting the feeding center and, 3)stimulating the satiety center in the hypothalamus. In Japan, mazindol is available for clinical use. We examined the effects of mazindol on 1) body weight, appetite, and abnormalities of obesity-related diseases in long-term use 2)maintenance of the reduced body weight after very-low-calorie diet (VLCD) therapy 3)combined use with VLCD therapy and, 4)inhibition of body weight gain in Prader-Willi syndrome. In long-term effects of mazindol, the average reduction of individual body weight was around 6.8 kg. The appetite of 59% of obese subjects was moderately suppressed. Systolic blood pressure, serum GOT, serum triglyceride, serum cholesterol, and glucose tolerance were also improved. With mazindol, 53.3% of obese subjects kept the reduced body weight after VLCD, in contrast, 20.0% of them kept it without mazindol. Combined use of mazindol with VLCD made the VLCD therapy more effective in out patients. Two of 3 patients with Prader-Willi syndrome inhibited their body weight gain with mazindol. Thus, mazindol produced positive effects in these studies, although the effects were limited.     

Identification of the Carboxyl-Terminal Processing Protease for the D1 Precursor Protein of the Photosystem II Reaction Center of Spinach

The enzyme involved in the carboxyl-terminal processing of theD1 precursor protein (pD1) of the photosystem II reaction centerwas purified from extracts of sonicated spinach thylakoids bya method that included chromatography on quaternary aminoethylanion-exchange, hydroxylapatite, copper-chelating affinity andgel-filtration columns. The enzyme was identified, from itschromatographic behavior, to be a monomeric protein of about45 kDa. The sequence of the amino-terminal 27 amino acids ofthis protein was determined directly, which exhibited low butappreciable (37%) homology to that deduced from a gene (ctpA)in Synechocystis sp. PCC 6803 that was proposed recently toencode the processing protease from results of genetic complementationanalysis. ³Present address: Asahi Kasei Chem. Co.     

An experimental model of ascending pyelonephritis due toCandida albicans in rats

We developed a new experimental model of ascendingCandida pyelonephritis in female rats with leukopenia and vesicoureteral reflux. Rats were treated transperitoneally with cyclophosphamide (200 mg/kg) to induce leukopenia 3 days before and transurethrally with diluted acetic acid solution to induce vesicoureteral reflux 1 day before inoculation ofCandida albicans strain, ATCC 10259 (containing 10⁷ cells). Microscopy revealed acute pyelonephritis in whichCandida cells invaded from the fornix and/or papilla into the medulla within 3 days after inoculation. Between 7 and 28 days after inoculation, chronic pyelonephritis reached the cortex. The incidence of pyelonephritis increased gradually and was approximately 80% after 7 days.Candida colony counts of bladder urine specimens obtained by direct puncture were significantly greater in rats with pyelonephritis extending into the parenchyma than in those with pyelonephritis located along the pelvis (p<0.01). These results suggest that this rat model shows the characteristic feature of ascending pyelonephritis due toC. albicans and that the severity ofCandida pyelonephritis can be estimated fromCandida counts of bladder urine.     

Detection of peptidoglycan and β-glucan with silkworm larvae plasma test

Abstract Discrimination of 70 Salmonella strains previously studied by ribotyping was realized by RAPD and ERIC-PCR analysis. RAPD results on the 56 S. typhimurium isolates did not closely match those of ribotyping. With ERIC-PCR, two fingerprints only were obtained. For the 14 S. enteritidis strains, a helpful discrimination was obtained with RAPD analysis, while ERIC-PCR resulted in a single fingerprint.