³¹P Saturation Transfer and Phosphocreatine Imaging in the Monkey Brain

³¹P magnetic resonance imaging with chemical-shift discrimination by selective excitation has been employed to determine the phosphocreatine (PCr) distribution in the brains of three juvenile macaque monkeys. PCr images were also obtained while saturating the resonance of the {gamma}-phosphate of ATP, which allowed the investigation of the chemical exchange between PCr and the {gamma}-phosphate of ATP catalyzed by creatine kinase. Superposition of the PCr images over the proton image of the same monkey brain revealed topological variations in the distribution of PCr and creatine kinase activity. PCr images were also obtained with and without visual stimulation. In two out of four experiments, an apparently localized decrease in PCr concentration was noted in visual cortex upon visual stimulation. This result is interpreted in terms of a possible role for the local ADP concentration in stimulating the accompanying metabolic response

Intensive Mutagenesis of the Nisin Hinge Leads to the Rational Design of Enhanced Derivatives

peer-reviewedNisin A is the most extensively studied lantibiotic and has been used as a preservative by the food industry since 1953. This 34 amino acid peptide contains three dehydrated amino acids and five thioether rings. These rings, resulting from one lanthionine and four methyllanthionine bridges, confer the peptide with its unique structure. Nisin A has two mechanisms of action, with the N-terminal domain of the peptide inhibiting cell wall synthesis through lipid II binding and the C-terminal domain responsible for pore-formation. The focus of this study is the three amino acid ‘hinge’ region (N 20, M 21 and K 22) which separates these two domains and allows for conformational flexibility. As all lantibiotics are gene encoded, novel variants can be generated through manipulation of the corresponding gene. A number of derivatives in which the hinge region was altered have previously been shown to possess enhanced antimicrobial activity. Here we take this approach further by employing simultaneous, indiscriminate site-saturation mutagenesis of all three hinge residues to create a novel bank of nisin derivative producers. Screening of this bank revealed that producers of peptides with hinge regions consisting of AAK, NAI and SLS displayed enhanced bioactivity against a variety of targets. These and other results suggested a preference for small, chiral amino acids within the hinge region, leading to the design and creation of producers of peptides with hinges consisting of AAA and SAA. These producers, and the corresponding peptides, exhibited enhanced bioactivity against Lactococcus lactis HP, Streptococcus agalactiae ATCC 13813, Mycobacterium smegmatis MC2155 and Staphylococcus aureus RF122 and thus represent the first example of nisin derivatives that possess enhanced activity as a consequence of rational design.This work was financed by a grant from the Irish Department of Agriculture, Food and the Marine through the Food Institutional Research Measure (08/RD/C/691) and with Science Foundation Investigator award (10/IN.1/B3027)

NMR in physiology and biomedicine, Robert J. Gillies, Ph.D. ed., Academic Press, San Diego, California, 1994. 471 pp. $95.00

No Abstract.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/38492/1/1910340121_ftp.pd

Thermal gravity, black holes and cosmological entropy

Taking seriously the interpretation of black hole entropy as the logarithm of the number of microstates, we argue that thermal gravitons may undergo a phase transition to a kind of black hole condensate. The phase transition proceeds via nucleation of black holes at a rate governed by a saddlepoint configuration whose free energy is of order the inverse temperature in Planck units. Whether the universe remains in a low entropy state as opposed to the high entropy black hole condensate depends sensitively on its thermal history. Our results may clarify an old observation of Penrose regarding the very low entropy state of the universe.Comment: 5 pages, 2 figures, RevTex. v4: to appear in Phys. Rev.

Mitochondrial Targeted Coenzyme Q, Superoxide, and Fuel Selectivity in Endothelial Cells

Background Previously, we reported that the “antioxidant” compound “mitoQ” (mitochondrial-targeted ubiquinol/ubiquinone) actually increased superoxide production by bovine aortic endothelial (BAE) cell mitochondria incubated with complex I but not complex II substrates. Methods and Results To further define the site of action of the targeted coenzyme Q compound, we extended these studies to include different substrate and inhibitor conditions. In addition, we assessed the effects of mitoquinone on mitochondrial respiration, measured respiration and mitochondrial membrane potential in intact cells, and tested the intriguing hypothesis that mitoquinone might impart fuel selectivity in intact BAE cells. In mitochondria respiring on differing concentrations of complex I substrates, mitoquinone and rotenone had interactive effects on ROS consistent with redox cycling at multiple sites within complex I. Mitoquinone increased respiration in isolated mitochondria respiring on complex I but not complex II substrates. Mitoquinone also increased oxygen consumption by intact BAE cells. Moreover, when added to intact cells at 50 to 1000 nM, mitoquinone increased glucose oxidation and reduced fat oxidation, at doses that did not alter membrane potential or induce cell toxicity. Although high dose mitoquinone reduced mitochondrial membrane potential, the positively charged mitochondrial-targeted cation, decyltriphenylphosphonium (mitoquinone without the coenzyme Q moiety), decreased membrane potential more than mitoquinone, but did not alter fuel selectivity. Therefore, non-specific effects of the positive charge were not responsible and the quinone moiety is required for altered nutrient selectivity. Conclusions In summary, the interactive effects of mitoquinone and rotenone are consistent with redox cycling at more than one site within complex I. In addition, mitoquinone has substrate dependent effects on mitochondrial respiration, increases repiration by intact cells, and alters fuel selectivity favoring glucose over fatty acid oxidation at the intact cell level.This work was supported by Veterans Affairs Medical Research Funds and grant DK25295 from the National Institutes of Health

Regional myocardial blood flow, function and metabolism using phosphorus-31 nuclear magnetic resonance spectroscopy during ischemia and reperfusion in dogs

Postreperfusion regional myocardial dysfunction may be associated with depletion of high energy phosphate compounds during ischemia and with their relatively slow repletion during reperfusion. However, few studies have correlated relatively rapid changes in regional myocardial function (sonomicrometers) and blood flow (microspheres) with high energy phosphate concentrations measured using phosphorus-31 nuclear magnetic resonance spectroscopy in intact large animal models of regional myocardial ischemia. The left anterior descending coronary artery of mongrel dogs was abruptly occluded for 17.1 ± 1.9 minutes and then completely released; measurements were made for an additional 22 minutes. Transmural blood flow decreased from 1.07 ± 0.25 to 0.25 ± 0.10 ml/(min × g) and holosystolic expansion was observed in all dogs (segmental systolic shortening decreased from 9.3 ± 3.7 to −6.3 ± 6.0%). Phosphocreatine (PCr) measured during 4.4 minute sampling intervals decreased to steady state within the first sampling period after occlusion and was 45.9 ± 17.0% of control at the end of the occlusion, whereas beta-adenosine triphosphate (beta-ATP) reached its lowest level early after reperfusion (72.7 ± 13.3% of control). The ratio of PCr to inorganic phosphate (Pi) decreased during the occlusion (3.34 ± 0.75 versus 1.01 ± 0.61) but returned to control level early during reperfusion. The ratio of PCr to beta-ATP also decreased during coronary occlusion (2.16 ± 0.39 versus 1.29 ± 0.39) but did not return to control level during reperfusion.Significant correlations were observed between the intensity of ischemia (reduced blood flow) and reductions in regional contractile function, PCr, beta-ATP, myocardial pH and the increase in Pi during the coronary occlusion. Also during ischemia, there were significant correlations between regional contractile function and both myocardial pH and Pi. PCr returned to control level rapidly after reperfusion (95.9 ± 13.2% of control in less than 5 minutes of reperfusion) whereas beta-ATP recovered only partially after 22 minutes (80.0 ± 17.5% of control). The correlation between the fraction of control beta-ATP and the fraction of control regional function at this time was r = 0.84 (p = 0.017).These results demonstrate metabolic correlates to regional myocardial ischemia in an intact dog model using phosphorus-31 spectroscopy. Additionally during reperfusion, beta-ATP, but not PCr, could be associated with the recovery of regional segmental contractile function