A temperature-sensitive Mycobacterium smegmatis glgE mutation leads to a loss of GlgE enzyme activity and thermostability and the accumulation of α-maltose-1-phosphate

Background: The bacterial GlgE pathway is the third known route to glycogen and is the only one present in mycobacteria. It contributes to the virulence of Mycobacterium tuberculosis. The involvement of GlgE in glycogen biosynthesis was discovered twenty years ago when the phenotype of a temperature-sensitive Mycobacterium smegmatis mutation was rescued by the glgE gene. The evidence at the time suggested glgE coded for a glucanase responsible for the hydrolysis of glycogen, in stark contrast with recent evidence showing GlgE to be a polymerase responsible for its biosynthesis. Methods: We reconstructed and examined the temperature-sensitive mutant and characterised the mutated GlgE enzyme. Results: The mutant strain accumulated the substrate for GlgE, α-maltose-1-phosphate, at the non-permissive temperature. The glycogen assay used in the original study was shown to give a false positive result with α-maltose-1-phosphate. The accumulation of α-maltose-1-phosphate was due to the lowering of the kcat of GlgE as well as a loss of stability 42 °C. The reported rescue of the phenotype by GarA could potentially involve an interaction with GlgE, but none was detected. Conclusions: We have been able to reconcile apparently contradictory observations and shed light on the basis for the phenotype of the temperature-sensitive mutation. General significance: This study highlights how the lowering of flux through the GlgE pathway can slow the growth mycobacteria

13C-direct detected NMR experiments for the sequential J-based resonance assignment of RNA oligonucleotides

We present here a set of 13C-direct detected NMR experiments to facilitate the resonance assignment of RNA oligonucleotides. Three experiments have been developed: (1) the (H)CC-TOCSY-experiment utilizing a virtual decoupling scheme to assign the intraresidual ribose 13C-spins, (2) the (H)CPC-experiment that correlates each phosphorus with the C4′ nuclei of adjacent nucleotides via J(C,P) couplings and (3) the (H)CPC-CCH-TOCSY-experiment that correlates the phosphorus nuclei with the respective C1′,H1′ ribose signals. The experiments were applied to two RNA hairpin structures. The current set of 13C-direct detected experiments allows direct and unambiguous assignment of the majority of the hetero nuclei and the identification of the individual ribose moieties following their sequential assignment. Thus, 13C-direct detected NMR methods constitute useful complements to the conventional 1H-detected approach for the resonance assignment of oligonucleotides that is often hindered by the limited chemical shift dispersion. The developed methods can also be applied to large deuterated RNAs

Physical activity and survival after a first myocardial infarction: The Corpus Christi Heart Project

Previous studies have demonstrated that habitual physical activity is associated with a reduced risk of incident coronary heart disease (CHD). However, the role of physical activity in lowering the risk of all-cause mortality, CHD mortality, reinfarction, or receipt of a revascularization procedure after a first myocardial infarction (MI) remains unresolved, particularly in minority populations. To investigate the associations between physical activity and risk of all-cause mortality, CHD mortality, reinfarction, and receipt of a revascularization procedure, this study was conducted among Mexican-American and non-Hispanic white women and men who survived a first MI. The Corpus Christi Heart Project, a population-based cardiovascular surveillance study, provide data which included vital status, survival time, medical history, CHD risk factor information, including level of physical activity among Mexican-American and non-Hispanic white adults who had experienced a first MI between May, 1988 and April, 1990. MI patients were interviewed at baseline and annually thereafter until their death or through May, 1995. A categorical variable was created to reflect change in level of physical activity following the first MI; categories included (1) sedentary with no change, (2) decreased activity, (3) increased activity, and (4) moderate activity with no change (the referent group). Proportional hazards regression analyses were used to assess the relationship of level of physical activity and risk of death, reinfarction, or receipt of a revascularization procedure adjusting for age, sex, ethnicity, severity of MI, and CHD risk factor status. Over a 7-year follow-up period, the relative risk (95% confidence intervals) of all-cause mortality was 4.67 (2.27, 9.60) for the sedentary-no change group, 2.33 (0.96, 5.67) for the decreased activity group, and 0.52 (0.11, 2.41) for the increased activity group. The relative risk of CHD mortality was 6.92 (2.05, 23.34) for the sedentary-no change group, 2.40 (0.55, 10.51) for the decreased activity group, and 1.58 (0.26, 9.65) for the increased activity group. The relative risk for reinfarction was 2.50 (1.52, 4.10) for the sedentary-no change group, 2.26 (1.24, 4.12) for the decreased activity group, and 0.52 (0.21, 1.32) for the increased activity group. Finally, the relative risk for receipt of a revascularization procedure was 0.65 (0.39, 1.07) for the sedentary-no change group, 0.45 (0.22, 0.92) for the decreased activity group, and 1.01 (0.51, 2.02) for the increased activity group. No interactions were observed for ethnicity or severity of first MI. These results are consistent with the hypothesis that moderate physical activity is independently associated with a lower risk of all-cause mortality, CHD mortality, and reinfarction, but not revascularization, among Mexican-American and non-Hispanic white, female and male, first MI patients. These results also support the current recommendation that physical activity plays an important role in the secondary prevention of CHD

Is a sedentary lifestyle an independent predictor for hospital and early mortality after elective cardiac surgery?

Contains fulltext : 119123.pdf (publisher's version ) (Open Access)OBJECTIVE: This study evaluates whether a sedentary lifestyle is an independent predictor for increased mortality after elective cardiac surgery. METHODS: Three thousand one hundred fifty patients undergoing elective cardiac surgery between January 2007 and June 2012 completed preoperatively the Corpus Christi Heart Project questionnaire concerning physical activity (PA). Based on this questionnaire, 1815 patients were classified as active and 1335 patients were classified as sedentary. The endpoints of the study were hospital mortality and early mortality. RESULTS: The study population had a mean age of 69.7 +/- 10.1 (19-95) years and a mean logistic EuroSCORE risk of 5.1 +/- 5.6 (0.88-73.8). Sedentary patients were significantly older (p = 0.001), obese (p = 0.001), had a higher EuroSCORE risk (p = 0.001), and a higher percentage of complications. Hospital mortality (1.1 % versus 0.4 % (p = 0.014)) and early mortality (1.5 % versus 0.6 % (p = 0.006)) were significantly higher in the sedentary group compared with the active group. However, a sedentary lifestyle was not identified as an independent predictor for hospital mortality (p = 0.61) or early mortality (p = 0.70). CONCLUSION: Sedentary patients were older, obese and had a higher EuroSCORE risk. They had significantly more postoperative complications, higher hospital mortality and early mortality. Despite these results, sedentary behaviour could not be identified as an independent predictor for hospital or early mortality