Carboxyatractylate-sensitive uncoupling in liver mitochondria from ground squirrels during hibernation and arousal

Energy coupling parameters of liver mitochondria from hibernating and arousing ground squirrels have been studied. In the oligomycin-treated mitochondria, carboxyatractylate, an inhibitor of the ATP/ADP-antiporter, is shown to decrease the respiration rate, to increase the membrane potential and to lower the rate of the membrane-potential discharge after the addition of cyanide to liver mitochondria from hibernating and arousing animals. BSA effectively substitutes for carboxyactactylate so that carboxyactactylate, added after BSA, has no effect. In mitochondria from hibernating animals, the maximal respiration rate in the presence of DNP and the rate of the membrane potential discharge in its absence are much lower than in those from arousing animals. It has been concluded that upon arousal of the animals from hibernation, the uncoupling of oxidative phosphorylation, mediated by free fatty acids and ATP/ADP-antiporter, parallels the respiratory chain activation.     

Proteins under extreme physical conditions

Life on earth is ubiquitous within the limits from -5 to 110 degrees C for temperature, 0.1 to 120 MPa for hydrostatic pressure, 1.0 to 0.6 for water activity and pH 1 to 12. In general, mutative adaptation of proteins to changing environmental conditions tends to maintain 'corresponding states' regarding overall topology, flexibility and hydration. Due to the minute changes in the free energy of stabilization responsible for enhanced stability, nature provides a wide variety of different adaptative strategies. In the case of thermophilic proteins, improved packing densities are crucial. In halophilic proteins, decreased hydrophobicity and clustered surface charges serve to increase water and salt binding required for solubilization at high salt concentration. In the case of barophiles, high-pressure adaptation is expected to be less important than adaptation to low temperatures governing the deep sea. Nothing is known with respect to the mechanisms underlying psychrophilic and acidophilic/alkalophilic adaptation.     

Neuraxin corresponds to a C-terminal fragment of microtubule-associated protein 5 (MAP5)

From cloned DNA, neuraxin has been identified as a tubulin binding protein of predicted molecular weight of 94 kDa. The deduced sequence of the rat protein exhibits high homology to the C-terminal region of mouse microtubule-associated protein 5 (MAP5). Here, we show that different neuraxin antibodies recognize MAP5, but fail to detect a protein of 94 kDa, in subcellular and microtubular fractions of the rat central nervous system. Furthermore, tubulin binding by neuraxin was found to be dependent on taxol. These data are consistent with neuraxin corresponding to a C-terminal fragment of MAP5 that contains a low-affinity tubulin binding site.     

Catabolism of naphthalene and salicylate byPseudomonas fluorescens

Analysis of spent naphthalene growth media ofPseudomonas fluorescens by GC-MS revealed the presence of salicylate. Gentisate 1,2-dioxygenase and pyrocatechol 1,2-dioxygenase were induced by growth on naphthalene, whereas only pyrocatechol 1,2-dioxygenase was induced during growth on salicylate. These results suggest the existence of alternative degradative routes of salicylate,via gentisate and pyrocatechol, which are involved in the catabolism of naphthalene.     

Novel UGA-suppressors in Escherichia coli K-12

UGA-specific nonsense suppressors from Escherichia coli K-12 were isolated and characterized. One of them (Su+UGA-11) was identified as a mutant of the prfB gene for the peptide releasing factor RF2. It appears that in this strain, while peptide release at sites of UGA mutations is retarded, the UGA stop codon is read through even in the absence of a tRNA suppressor, exhibiting a novel type of passive nonsense suppression. Three suppressors (Su+UGA-12, -16 and -34) were capable of restoring the streptomycin sensitive phenotype in resistant bacteria (strAr). Because of their drug-related phenotype, these are possibly mutations in the components of the ribosomal machinery, particularly those concerned with peptide release at UGA nonsense codons. A tRNA suppressor was also obtained which was derived from the tRNA(Trp) gene. In this strain, a long region between rrnC (84.5 min) and rrnB (89.5 min) was duplicated and one of the duplicated genes of tRNA(Trp) was mutated to the suppressor. The mechanism of UGA-suppression is discussed in terms of translation termination at the nonsense codon in both active and passive fashions.     

Ion selectivity of the Vibrio alginolyticus flagellar motor.

The marine bacterium, Vibrio alginolyticus, normally requires sodium for motility. We found that lithium will substitute for sodium. In neutral pH buffers, the membrane potential and swimming speed of glycolyzing bacteria reached maximal values as sodium or lithium concentration was increased. While the maximal potentials obtained in the two cations were comparable, the maximal swimming speed was substantially lower in lithium. Over a wide range of sodium concentration, the bacteria maintained an invariant sodium electrochemical potential as determined by membrane potential and intracellular sodium measurements. Over this range the increase of swimming speed took Michaelis-Menten form. Artificial energization of swimming motility required imposition of a voltage difference in concert with a sodium pulse. The cation selectivity and concentration dependence exhibited by the motile apparatus depended on the viscosity of the medium. In high-viscosity media, swimming speeds were relatively independent of either ion type or concentration. These facts parallel and extend observations of the swimming behavior of bacteria propelled by proton-powered flagella. In particular, they show that ion transfers limit unloaded motor speed in this bacterium and imply that the coupling between ion transfers and force generation must be fairly tight.     

Physiological implications of the substrate specificities of acetohydroxy acid synthases from varied organisms.

Acetohydroxy acid synthase (AHAS; EC 4.1.3.18) catalyzes the following two parallel, physiologically important reactions: condensation of two molecules of pyruvate to form acetolactate (AL), in the pathway to valine and leucine, and condensation of pyruvate plus 2-ketobutyrate to form acetohydroxybutyrate (AHB), in the pathway to isoleucine. We have determined the specificity ratio R with regard to these two reactions (where VAHB and VAL are rates of formation of the respective products) as follows: VAHB/VAL = R [2-ketobutyrate]/[pyruvate] for 14 enzymes from 10 procaryotic and eucaryotic organisms. Each organism considered has at least one AHAS of R greater than 20, and some appear to contain but a single biosynthetic AHAS. The implications of this for the design of the pathway are discussed. The selective pressure for high specificity for 2-ketobutyrate versus pyruvate implies that the 2-ketobutyrate concentration is much lower than the pyruvate concentration in all these organisms. It seems important for 2-ketobutyrate levels to be relatively low to avoid a variety of metabolic interferences. These results also reinforce the conclusion that biosynthetic AHAS isozymes of low R (1 to 2) are a special adaptation for heterotrophic growth on certain poor carbon sources. Two catabolic "pH 6 AL-synthesizing enzymes" are shown to be highly specific for AL formation only (R less than 0.1).     

Transbilayer distribution and mobility of phosphatidylinositol in human red blood cells

The present studies describe the distribution of phosphatidylinositol (PI) within the membrane bilayer of the human red blood cell (RBC) as well as its transbilayer mobility. The membrane bilayer distribution was determined by measuring the hydrolysis of PI in the exterior leaflet of the RBC membrane using a PI-specific phospholipase C and by extraction of PI from the exterior leaflet using bovine serum albumin. The transbilayer mobility of PI was measured by following the fate of radiolabeled PI which was first incorporated into the outer leaflet of the RBC membrane. Our results indicate that PI is asymmetrically distributed in the membrane, with approximately 80% located in the inner and 20% in the outer leaflet of the bilayer. The rate of transbilayer mobility of PI is similar to that for certain molecular species of phosphatidylcholine and much slower than that reported for the aminophospholipids in the RBC membrane.     

Vectorial secretion of extracellular matrix proteins, matrix-degrading proteinases, and tissue inhibitor of metalloproteinases by endothelial cells

In a study of the vectorial secretion of proteins by bovine aortic arch endothelial cells, we found that the extracellular matrix macromolecules collagen and fibronectin as well as several matrix-degrading metalloproteinases were secreted selectively in the basal direction. In contrast, the tissue inhibitor of metalloproteinases showed only a weak preference for the basal direction. Three proteins at 18-35 kDa were secreted with preference apically, counter to the basal secretion of approximately 70% of the total secreted protein. As expected, rabbit synovial fibroblasts, which were used as a control, secreted proteins, including collagen, gelatin-degrading proteinases, and casein-degrading proteinases, equally in apical and basal directions. The basal secretion of collagen, fibronectin, gelatinases, and tissue inhibitor of metalloproteinases by bovine aortic arch endothelial cells suggests that the structural and functional polarity of these cells is manifested, in part, at the level of polarized secretion of matrix-related proteins.     

Morphofunctional status of the system of mononuclear phagocytes in rats exposed to the embryotoxic effect of cadmium

The experiments have been performed on white non-inbred rats, subjected to intragastric injection of cadmium chloride (7.5 mg/kg) during the whole period of pregnancy. Morphological analysis has been performed on the state of the main links in mononuclear phagocytes system (osseous-medullary precursors, monocytes of blood and alveolar macrophages of lungs), using cytological, cytochemical and electron microscopical (scanning electron microscopy) investigation methods. Changes in the cell enzymatic activity, inhibition of adhesiveness and lesion of the superficial architectonics of blood monocytes and pulmonary macrophages have been revealed against the background of their increased phagocyting activity. This demonstrates certain decrease in nonspecific resistivity of the organism, when the embryotoxic effect is developing.