Application of the tryptophanase promoter to high expression of the tryptophan synthase gene inEscherichia coli

Summary The application of an inducible regulation system using the trytophanase operon promoter (TPase promoter; P_tna) was examined for its high expression of the tryptophan synthase (TS) gene in Escherichia coli. The main problem in the application of P_tna for industrial purposes is catabolite repression by glucose, since glucose is the most abundant carbon source. However, this problem could be avoided by changing glucose to an organic acid, such as succinate, fumarate, malate and acetate, in the course of cultivation after glucose initially added was completely consumed. Under these conditions, l-tryptophan was also used to induce tryptophan synthase. Thus, the specific activity of TS in E. coli strain no. 168 harbouring pBR322F-P_tnaTS was increased 500-fold compared to that of the cultured host strain. About 1 mol l-tryptophan/l reaction mixture was formed from indole and l-serine at 37° C for 3.5 h. Offprint requests to: H. Yukawa     

Comparison of muscle force,muscle endurance,and electromyogram activity during an expedition at high altitude

Handgrip force (HF), maximal pinch force (MF), muscle endurance (ME), and the median power frequency (MdPF) of the activity shown in the electromyogram (EMG) were studied at various altitudes in eight normal healthy subjects. MF and ME were measured between the index finger and thumb, and all measurements were obtained at altitudes ranging from 610 to 4860 m during an expedition in the Qinghai Plateau in China. With the change in altitude HF, ME, and MF showed no significant change. Compared to the MdPF at 2260 m on ascent, the MdPF at other altitudes showed a significant decrease (P<0.01). Thus, we conclude that muscle performance (HF, MF, and ME) was not affected by the environment at high altitude. However, MdPF was affected and the mean MdPF at 610 m after the expedition did not recover to initial values of MdPF. We suggest these results may have been affected by fatigue and chronic exposure to the hypobaric hypoxic environment, since the members of the expedition party expressed feelings of sluggishness and fatigue after the expedition.     

Cardiopulmonary function in bicycle racing over mountainous terrain at moderate altitude

To examine cardiopulmonary function during exercise in a mountainous region at moderate altitude, we measured cardiac frequency, oxygen consumption , and percentage arterial hemoglobin oxygen saturation (%SaO₂) before and after a bicycle race with a starting point at 638 m and finishing point at 1980 m. The time required to ascend an elevation of 10 m was prolonged with increasing altitude, and heart rate also increased with altitude. The %SaO₂ at the starting point and at the finishing point differed significantly (P<0.01). Faster cyclists exhibited higher %SaO₂ and lower , while slower cyclists exhibited a reduction in %SaO₂ and an increase in immediately after the race. The %SaO₂ recovery time was significantly correlated with the racing time (r=0.54,P<0.001). Therefore, the faster cyclists' oxygen debt upon completion of the race may be small and recovery of cardiopulmonary function may be fast, while the slower cyclists' oxygen debt may be large and recovery of cardiopulmonary function may be slow.     

Neurocalcin: a novel calcium-binding protein from bovine brain.

A novel calcium-binding protein (molecular weight 23,000-24,000, pI 5.3-5.5), which we term neurocalcin, was identified in bovine brain. Using calcium-dependent drug affinity chromatography ((S)-P-(2-aminoethyloxy)-N-[2-(4-benzyloxycarbonylpiperazinyl++ +)-1-(P- methoxybenzyl)ethyl]-N-methylbenzene-sulfonamide dihydrochloride, W-77, -coupled Sepharose 6B), we purified neurocalcin from bovine brain. The partial amino acid sequence of neurocalcin revealed it to be an as yet unidentified protein with three putative calcium binding sites (EF-hands). Further purification and sequence analysis demonstrated the presence of four isoprotein forms designated alpha, beta, gamma 1, and gamma 2. When the 165 sequenced residues of neurocalcin beta are compared with sequences of other proteins, neurocalcin beta has a 38.2% sequence homology with visinin and 45.5% with recoverin (Yamagata, K., Goto, K., Kuo, C.-H., Kondo, H., and Miki, N. (1990) Neuron 2, 469-476; Dizhoor, A. M., Ray, S., Kumar, S., Niemi, G., Spencer, M., Brolley, D., Walsh, K. A., Philipov, P. P., Hurley, J. B., and Stryer, L. (1991) Science 251, 915-918). Both visinin and recoverin are expressed specifically in retinal photoreceptors and are not found in brain. Unlike visinin and recoverin, neurocalcin is purified not only from retina but also from bovine brain. Our results suggest that neurocalcin is a recoverin-like protein expressed in bovine brain.     

Testosterone potentiation of the effectiveness of ACTH1-24 on the induction of the stretch-yawning syndrome (SYS) in male guinea pigs

Ventricular administration of ACTH^1–24 stimulated stretch-yawning in castrated male guinea pigs in a dose-related manner. Daily subcutaneous treatment with testosterone propionate (TP) facilitated the effects of ACTH^1–24 on this response. TP given without ACTH^1–24 stimulated yawning, but not stretching or combined stretch-yawning. Unlike most other species, guinea pigs did not display auto-grooming, scratching, or wet-dog shaking in response to intracranial ACTH^1–24. The results suggest that testosterone may alter the sensitivity of neural mechanisms which are responsive to ACTH^1–24.     

Partial silencing of fucosyltransferase 8 gene expression inhibits proliferation of Ishikawa cells,a cell line of endometrial cancer

Endometrial cancer is the most common gynecologic malignancy and is associated with increased morbidity each year, including young people. However, its mechanisms of proliferation and progression are not fully elucidated. It is well known that abnormal glycosylation is involved in oncogenesis, and fucosylation is one of the most important types of glycosylation. In particular, fucosyltransferase 8 (FUT8) is the only FUT responsible for α1, 6-linked fucosylation (core fucosylation), and it is involved in various physiological as well as pathophysiological processes, including cancer biology. Therefore, we aimed to identify the expression of FUT8 in endometrial endometrioid carcinoma and investigate the effect of the partial silencing of the FUT8 gene on the cell proliferation of Ishikawa cells, an epithelial-like endometrial cancer cell line. Quantitative real-time PCR analysis showed that FUT8 gene expression was significantly elevated in the endometrial endometrioid carcinoma, compared to the normal endometrium. The immunostaining of FUT8 and Ulex europaeus Agglutinin 1 (UEA-1), a kind of lectin family specifically binding to fucose, was detected endometrial endometrioid carcinoma. The proliferation assay showed FUT8 partial knockdown by transfection of siRNA significantly suppressed the proliferation of Ishikawa cells, concomitant with the upregulation in the gene expressions associated with the interesting pathways associated with de-ubiquitination, aspirin trigger, mesenchymal-epithelial transition (MET) et al. It was suggested that the core fucosylation brought about by FUT8 might be involved in the proliferation of endometrial endometrioid carcinoma cells.     

3H-31, A Non-structural Protein of Heliothis virescens ascovirus 3h,Inhibits the Host Larval Cathepsin and Chitinase Activities

Virologica Sinica - 3h-31 of Heliothis virescens ascovirus 3h (HvAV-3h) is a highly conserved gene of ascoviruses. As an early gene of HvAV-3h, 3h-31 codes for a non-structural protein (3H-31) of...