Reconstitution and transphosphorylation of TGF-beta receptor complexes.

Transforming growth factor-beta (TGF-beta) signals by contacting two distantly related transmembrane serine/threonine kinases called receptors I (T beta R-I) and II (T beta R-II). TGF-beta binds to T beta R-II, which is a constitutively active kinase and this complex recruits T beta R-I, causing its phosphorylation and signal propagation to downstream substrates. The biochemical properties of this interaction were analyzed with reconstituted receptor systems. T beta R-I and T beta R-II baculovirally expressed at high levels in insect cells have the ligand binding properties of receptors expressed in mammalian cells, and form a complex in which T beta R-I phosphorylation is dependent on the kinase activity of T beta R-II. Furthermore, T beta R-I and T beta R-II can form a complex in vitro, and their cytoplasmic domains can specifically interact in a yeast two-hybrid system. In vitro complex formation with catalytically active T beta R-II is necessary and sufficient for T beta R-I phosphorylation, which within this complex does not require the catalytic activity of T beta R-I, thus mimicking T beta R-I phosphorylation in intact cells. In addition, T beta R-I phosphorylated in vitro remains associated with T beta R-II. These results suggest that T beta R-I and T beta R-II have affinity for each other, however, the ligand is required for stable complex formation under physiological conditions. Once formed, this complex is sufficient for T beta R-I phosphorylation by T beta R-II.     

Effect of prior ingestion of glucose or fructose on the performance of exercise of intermediate duration

The metabolic responses induced by the ingestion of a beverage containing glucose (G), fructose (F) or placebo (W) 30 min before exercise of high intensity and intermediate duration have been investigated; in these conditions the energy processes are mostly dependent on aerobic reactions. A group of 11 male recreational sportsmen ran on a treadmill, at an intensity corresponding to 82% of peak oxygen consumption, until exhaustion on three different occasions (after ingestion of a beverage containing 75 g of G, 75 g of F or W). Plasma glucose, insulin, and lactic acid concentrations were determined just prior to the ingestion of the beverages, 30 min afterwards and 10 and 30 min after completion of the exercise. The mean endurance time was 644 (SD 261) s after the ingestion of G, 611 (SD 227) s after the ingestion of F and 584 (SD 189) s after the ingestion of the W (P < 0.05 between G and W). No differences in the oxygen uptake, respiratory quotient or lactate concentrations between the three trials were observed. Both plasma glucose and insulin concentrations determined in samples obtained immediately before the onset of exercise were higher when G was ingested than when F (P < 0.05 andP < 0.05, respectively) or W (P < 0.001 and P < 0.005, respectively) were ingested. These findings would suggest that the ingestion of G prior to an effort of intermediate duration may improve physical performance.     

Evolution of the ATP-binding-cassette transmembrane transporters of vertebrates

The ATP-binding-cassette transmembrane transporters (ABC transporters) known from vertebrates belong to four major subfamilies: (1) the P- glycoproteins (Pgp); (2) the cystic fibrosis transmembrane conductance regulators (CFTR); (3) the Tap proteins encoded with the major histocompatibility complex of mammals; and (4) the peroxisomal membrane proteins. Both Pgp and CFTR have a structure suggesting a past internal gene duplication; a phylogenetic analysis indicated that these duplications occurred independently, while an independent tandem gene duplication occurred in the case of the Tap family. Both the Pgp and Tap proteins show evidence of relationship to bacterial ABC transporters lacking internal duplication, and both are significantly more closely related to the HlyB and MsbA families of transporters from purple bacteria than they are to ABC transporters from nonpurple bacteria. The simplest hypothesis to explain this observation is that eukaryotic Pgp and Tap genes are descended from a mitochondrial gene or genes that were subsequently translocated to the nuclear genome. The Pgp genes of eukaryotes are characterized by a remarkable degree of convergent evolution between the ATP-binding cassettes of their N- terminal and C-terminal halves, whereas no such convergence is seen between the two halves of CFTR genes or between the duplicated Tap genes. Exon 13 of the CFTR gene, which encodes a putative regulatory domain not found in other ABC transporters apart from CFTR, showed high levels of both synonymous and nonsynonymous difference in comparisons among different mammalian species, suggesting that this region is a mutational hot spot.      

Protein stabilization by compatible solutes. Effect of diglycerol phosphate on the dynamics of Desulfovibrio gigas rubredoxin studied by NMR.

Heteronuclear NMR relaxation measurements and hydrogen exchange data have been used to characterize protein dynamics in the presence or absence of stabilizing solutes from hyperthermophiles. Rubredoxin from Desulfovibrio gigas was selected as a model protein and the effect of diglycerol phosphate on its dynamic behaviour was studied. The presence of 100 mM diglycerol phosphate induces a fourfold increase in the half-life for thermal denaturation of D. gigas rubredoxin. A model-free analysis of the protein backbone relaxation parameters shows an average increase of generalized order parameters of 0.015 reflecting a small overall reduction in mobility of fast-scale motions. Hydrogen exchange data acquired over a temperature span of 20 degrees C yielded thermodynamic parameters for the structural opening reactions that allow for the exchange. This shows that the closed form of the protein is stabilized by an additional 1.6 kJ x mol(-1) in the presence of the solute. The results seem to indicate that the stabilizing effect is due mainly to a reduction in mobility of the slower, larger-scale motions within the protein structure with an associated increase in the enthalpy of interactions.     

Evidence for the presence of β-lactamase inStreptomyces glaucescens and its inhibition by sodium clavulanate

Streptomyces glaucescens is shown to possess -lactamase activity which is inhibitable by clavulanate. This is important in regard to its use as a cloning host for enzymes of \-lactam biosynthesis.     

Urinary excretion of PGE2 and renal function in acute myocardial infarction

In 17 hospitalized patients affected by acute myocardial infarction (AMI) PGE2 urinary excretion, renal function and, furthermore, cortisol urinary excretion were tested during a 21 days trial. In 12 patients all the parameters under consideration underwent a similar trend: PGE2 urinary excretion exactly like glomerular filtration rate, Na+ excretion and diuresis tended to be reduced during the first 5 days and they rapidly recovered the normality after this period. Cortisol urinary excretion displayed a characteristic pattern: i.e. the highest values were observed in the first days, followed by a progressive decrease towards physiological levels since the 4th day. Different findings were obtained in 5 cases treated with an antiinflammatory drug (Indoprophen i.m. 200 mg x2 die). In fact the low levels of urinary PGE2 on the first days did not display any increasing and GFR, urinary flow, and Na+ tubular balance underwent irregular and not significant variations. These data suggest that an impaired Prostaglandin synthesis may be related to a compromised renal function often occurring in AMI.     

Structural and functional properties of rat liver mitochondria in hexachlorobenzene induced experimental porphyria

A possible link between changes in iron and porphyrin content in liver mitochondria, from rats treated with either hexachlorobenzene, iron, or hexachlorobenzene plus iron, as a function of treatment time and their structural-functional properties, has been investigated. Normal oxidative phosphorylation in mitochondria from rats treated with iron has been shown. By contrast a significant and constant uncoupling of the phosphorylative process, fully reversed by albumin, in mitochondria from rats treated with hexachlorobenzene and hexachlorobenzene plus iron has been presented. A possible involvement of pentachlorophenol in causing these abnormalities has been proposed.