'98 Escherichia coli SWISS-2DPAGE database update

The combination of two-dimensional polyacrylamide gel electrophoresis (2-D PAGE), computer image analysis and several protein identification techniques allowed the Escherichia coli SWISS-2DPAGE database to be established. This is part of the ExPASy molecular biology server accessible through the WWW at the URL address http://www.expasy.ch/ch2d/ch2d-top.html . Here we report recent progress in the development of the E. coli SWISS-2DPAGE database. Proteins were separated with immobilized pH gradients in the first dimension and sodium dodecyl sulfate-polyacrylamide gel electrophoresis in the second dimension. To increase the resolution of the separation and thus the number of identified proteins, a variety of wide and narrow range immobilized pH gradients were used in the first dimension. Micropreparative gels were electroblotted onto polyvinylidene difluoride membranes and spots were visualized by amido black staining. Protein identification techniques such as amino acid composition analysis, gel comparison and microsequencing were used, as well as a recently described Edman "sequence tag" approach. Some of the above identification techniques were coupled with database searching tools. Currently 231 polypeptides are identified on the E. coli SWISS-2DPAGE map: 64 have been identified by N-terminal microsequencing, 39 by amino acid composition, and 82 by sequence tag. Of 153 proteins putatively identified by gel comparison, 65 have been confirmed. Many proteins have been identified using more than one technique. Faster progress in the E. coli proteome project will now be possible with advances in biochemical methodology and with the completion of the entire E. coli genome.     

Addressing the tertiary structure of human parathyroid hormone-(1-34)

Parathyroid hormone (PTH) regulates mineral metabolism and bone turnover by activating specific receptors located on osteoblastic and renal tubular cells and is fully functional as the N-terminal 1-34 fragment, PTH-(1-34). Previously, a "U-shaped" conformation with N- and C-terminal helices brought in close proximity by a turn has been postulated. The general acceptance of this hypothesis, despite limited experimental evidence, has altered the direction of the design of PTH-analogs. Examining the structure of human PTH-(1-34) under conditions that encompass the different environments the hormone may experience in the approach to and interaction with the G-protein-coupled receptor (including benign aqueous and saline solutions and in the presence of dodecylphosphocholine), we observe no evidence for a U-shape conformation or any tertiary structure. Instead, the N- and C-terminal helical domains, which vary in length and stability depending on the conditions, are separated by a highly flexible region of undefined conformation. These observations are in complete accord with recent conformational studies of PTH-related protein analogs containing lactams (Mierke, D. F., Maretto, S., Schievano, E. , DeLuca, D., Bisello, A., Mammi, S., Rosenblatt, M., Peggion, E., and Chorev, M. (1997) Biochemistry 36, 10372-10383) or a model amphiphilic alpha-helix (Pellegrini, M., Bisello, A., Rosenblatt, M., Chorev, M., and Mierke, D. F. (1997) J. Med. Chem. 40, 3025-3031). Reliable structural data from different environmental conditions are absolutely requisite for the next step in the design of non-peptide PTH analogs.     

Accuracy and efficacy of chest radiography in the intensive care unit

264 patients with cancer of larynx, 21 female and 234 male, had a testosterone and sex hormone binding globulin (SHBG) before the treatment in serum estimated. Because of dependence of levels of hormones the group of patients was divided into three following groups: "lower than standard", "standard", "higher than standard". The correlation between these groups and sex, age, localization of tumor, organs' advances, stage of morphological malignancy and type of cancer was reported. Anomalous values of testosterone were in male group more frequently reported. Anomalous values of SHBG were similar in male and female groups, but in the female group there was a significant majority of "lower than standard" values reported. The majority of abnormal values of testosterone and SHBG was described in groups of age higher than 50 years. There were no differences in testosterone and SHBG levels in different localization of tumors in larynx. In advanced stage T3 and T4 there were more frequent lower mean values of testosterone levels and higher values of SHBG levels in comparison to T2 stages. In tumors in G1 and G2 stages of histological malignancy higher levels of SHBG and higher mean levels of testosterone. The tumors in stage G3 the hormone levels were lowers were observed. The levels of SHBG in groups of carcinoma planoepitheliale keratodes were in 66% higher than in a group of carcinoma planoepitheliale akeratodes but in both groups the levels of testosterone were nearing the same. In group of patients with larynx cancer the negative correlation between the levels of testosterone and SHBG was not observed. Higher SHBG levels were not always accompanied by lower testosterone levels.     

Cortisol in physiological concentrations acts within minutes to modify effects of prolactin and growth hormone on prostaglandin secretion: importance of effect in modulating cellular responses to calcium and cyclic nucleotides

We propose that intracellular prostaglandins (PGs) are essential for the final expression of the effects of second messengers in most cells. We suggest that the amounts of PGs required are very small (in the picomolar range) and are much lower than those used in most current PG studies. We suggest that while therapeutic levels of inhibitors of PG synthetase may be adequate to block the overflow of PGs from cells, they are in most cases unlikely to reduce intracellular PGs sufficiently to test the role of such PGs. We propose that there is a basal level of PG synthesis unaffected by hormones but that above this level PG synthesis is regulated by the interplay between physiological levels of cortisol, prolactin, growth hormone and thyroid hormones. For the most part prolactin seems to stimulate PG synthesis and cortisol to inhibit it: cortisol has, however, no inhibitory effect on basal PG synthesis. In reducing prolactin-stimulated PG synthesis cortisol is 1000-2000 times more potent than indomethacin on a molar basis. We suggest that the regulation of intracellular PG levels is the mechanism of the so-called "permissive" actions of these hormones. These concepts could prove important in the understanding of many aspects of physiology and pathophysiology including diurnal and seasonal changes in hormone responsiveness. They are also relevant to the use of established drugs and the design of new ones.     

Leukemic reticuloendotheliosis: a medical technologist's diagnosis

Leukemic reticuloendotheliosis (LRE) or "hairy cell leukemia" is a unique form of leukemia which possesses features of both a chronic and acute leukemia. Its characteristic morphology, clinical diagnosis, and treatment are discussed. A high index of suspicion should make this entity easily diagnosed by the medical technologist.     

Effect of the chemical structure of aromatic polyimides on their thermal aging,relaxation behavior and mechanical properties

This article examines the effects of structural changes and thermal aging treatments on the relaxation processes and mechanical properties of three polyimides differing for their molecular structure i.e. PMDA-ODA, 6FDA-ODA, and 6FDA-6FpDA. These polyimides were obtained by thermal imidization of their polyamic acid precursors, which were synthesized from the respective dianhydrides [pyromellitic anhydride (PMDA), hexahydrofluoroisopropylidene diphthalic anhydride (6FDA)], and diamines [4,4′-diaminodiphenyl ether (ODA), 4,4′-(hexafluoroisopropylidene) dianiline (6FpDA)]. After the curing process, the polyimides were thermally aged at a fixed temperature for various times Dynamic mechanical measurements performed in a multi-frequency mode, were used to determine the glass-rubber and sub-glass transitions, as well as the activation energy of the β transition. It was found that the T _g decreased in the order PMDA-ODA > 6FDA-6FpDA > 6FDA-ODA as a result of an increased chain rigidity and molecular packing induced by charge transfer interactions during the thermal imidization process. The β sub-glass transition showed two relaxation processes identified as β′ and β′′. The β′ process was attributed to the local motion of the diamine constituents while the β′′ process was caused by the local motion of the dianhydride moiety. The cooperativity of these molecular motions was also assessed via the Starkweather method. The thermal aging enhanced the state of aggregation of polyimide chains and thus the T _g and the sub-glass transition properties. This effect was particularly marked for the PMDA-ODA polyimide. Also the mechanical properties were significantly affected by chemical structure and aging treatments. For non-aged samples the more influenced parameter was the elongation at break, which decreased in the order PMDA-ODA > 6FDA-ODA > 6FDA-6FpDA. The aging enhanced the elastic modulus and the tensile strength and reduced the elongation at break.     

Growth performance,nutrient digestibility and immune response of broiler chicks fed diets supplemented with a culture of Lactobacillus bulgaricus

BACKGROUND: Probiotics are being developed for use in animal feed to enhance production performance and prevention of gastrointestinal infections. The ban on using antibiotics as growth promoters, antibiotic resistance and the inherent problems of developing new vaccines make a compelling case for developing alternatives for in‐feed antibiotics. The alternatives of choice have to be considered under the environmental conditions of the animal. Among the probiotics in use today, Lactobacillus has been shown to play a vital role in disease prevention, immune enhancement, improved growth and carcass yield in poultry. The present study investigates the effect of Lactobacillus bulgaricus (LB)‐based probiotic on the growth performance, nutrient digestibility and immune response of broilers under tropical environmental conditions. RESULTS: Broilers fed LB diets consumed more feed (P < 0.05) and had greater body weight gain than the control group. Feed/gain ratio improved significantly (P < 0.05) with the 20, 40 and 60 mg kg^?1 LB diets compared with the control or 80 mg kg^?1 LB diet. The apparent digestibilities of nitrogen and fat increased with LB supplementation. However, there was no significant difference (P > 0.05) in fibre digestibility. White blood cell count increased significantly in broilers fed higher levels (>40 mg kg^?1) of LB compared with the control group. Antibody production measured as antibody titre against Newcastle disease vaccine showed a curvilinear response over the range of LB concentrations examined. CONCLUSION: The results indicate that LB addition to broiler chick diets significantly improved growth performance, increased nutrient digestibility and stimulated humoral immune response. Copyright © 2008 Society of Chemical Industry     

Dominant effects of the bcr-abl oncogene on Drosophila morphogenesis

The Ras protein and its homolog, Rap1A, have an identical "effector region" (residues 32-40) preceded by Asp30-Glu31 and Glu30-Lys31, respectively. In the complex of the "Ras-like" E30D/K31E mutant Rap1A with the Ras-binding domain (RBD), residues 51-131 of Raf-1, Glu31 in Rap1A forms a tight salt bridge with Lys84 in Raf-1. However, we have recently found that Raf-1 RBD binding of Ras is indeed reduced by the E31K mutation, but is not affected by the E31A mutation. Here, the "Rap1A-like" D30E/E31K mutant of Ras was prepared and shown to bind the Raf-1 RBD less strongly than wild-type Ras, but slightly more tightly than the E31K mutant. The backbone 1H, 13C, and 15N magnetic resonances of the Raf-1 RBD were assigned in complexes with the wild-type and D30E/E31K mutant Ras proteins in the guanosine 5'-O-(beta,gamma-imidotriphosphate)-bound form. The Lys84 residue in the Raf-1 RBD exhibited a large change in chemical shift upon binding wild-type Ras, suggesting that Lys84 interacts with wild-type Ras. The D30E/E31K mutant of Ras caused nearly the same perturbations in Raf-1 chemical shifts, including that of Lys84. We hypothesized that Glu31 in Ras may not be the major salt bridge partner of Lys84 in Raf-1. A molecular dynamics simulation of a model structure of the Raf-1 RBD.Ras.GTP complex suggested that Lys84 in Raf-1 might instead form a tight salt bridge with Asp33 in Ras. Consistent with this, the D33A mutation in Ras greatly reduced its Raf-I RBD binding activity. We conclude that the major salt bridge partner of Lys84 in Raf-1 may be Asp33 in Ras.