Human Semaphorin 3 Variants Link Melanocortin Circuit Development and Energy Balance

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Mechanistic aspects of the covalent flavoprotein dimethylglycine oxidase of Arthrobacter globiformis studied by stopped-flow spectrophotometry

Dimethylglycine oxidase (DMGO) is a covalent flavoenzyme from Arthrobacter globiformis that catalyzes the oxidative demethylation of dimethylglycine to yield sarcosine, formaldehyde, and hydrogen peroxide. Stopped-flow and steady-state kinetic studies have been used to study the reductive and oxidative half-reactions using dimethylglycine and O2 as substrates. The reductive half-reaction is triphasic. The rate of the fast phase is dependent on substrate concentration, involves flavin reduction, and has a limiting rate constant of 244 s(-1). This phase also displays a kinetic isotope effect of 2.9. Completion of the first kinetic phase generates an intermediate with broad spectral signature between 350 and 500 nm, which is attributed to a reduced enzyme-iminium charge-transfer species, similar to the purple intermediate that accumulates in reactions of D-amino acid oxidase (DAAO) with alanine. The second phase (16 s(-1)) is independent of substrate concentration and is attributed to iminium hydrolysis/deprotonation. The third phase (2 s(-1)) is attributed to product release, the rate of which is less than the steady-state turnover rate (10.6 s(-1)). Flavin oxidation of dithionite- and dimethylglycine-reduced enzyme by O2 occurs in a single phase, and the rate shows a linear dependence on oxygen concentration, giving bimolecular rate constants of 342 and 201 mM(-1) x s(-1), respectively. Enzyme-monitored turnover experiments indicate that decay of the reduced enzyme-iminium intermediate is rate-limiting, consistent with rate constants determined from single turnover studies. A minimal kinetic mechanism is presented, which establishes a close relationship to the mechanism of action of DAAO. The covalent flavin in dimethylglycine oxidase is identified as an alphaN1-histidyl48-FAD, and equilibrium titration studies establish a single redox center that displays typical flavoprotein 'oxidase' characteristics.     

Predictors of mortality of patients with acute respiratory failure secondary to chronic obstructive pulmonary disease admitted to an intensive care unit: A one year study

Background

Patients with acute exacerbation of chronic obstructive pulmonary disease (COPD) commonly require hospitalization and admission to intensive care unit (ICU). It is useful to identify patients at the time of admission who are likely to have poor outcome. This study was carried out to define the predictors of mortality in patients with acute exacerbation of COPD and to device a scoring system using the baseline physiological variables for prognosticating these patients.

Methods

Eighty-two patients with acute respiratory failure secondary to COPD admitted to medical ICU over a one-year period were included. Clinical and demographic profile at the time of admission to ICU including APACHE II score and Glasgow coma scale were recorded at the time of admission to ICU. In addition, acid base disorders, renal functions, liver functions and serum albumin, were recorded at the time of presentation. Primary outcome measure was hospital mortality.

Results

Invasive ventilation was required in 69 patients (84.1%). Fifty-two patients survived to hospital discharge (63.4%). APACHE II score at the time of admission to ICU {odds ratio (95 % CI): 1.32 (1.138–1.532); p < 0.001} and serum albumin (done within 24 hours of admission) {odds ratio (95 % CI): 0.114 (0.03-0.432); p = 0.001}. An equation, constructed using the adjusted odds ratio for the two parameters, had an area under the ROC curve of 91.3%. For the choice of cut-off, sensitivity, specificity, positive and negative predictive value for predicting outcome was 90%, 86.5%, 79.4% and 93.7%.

Conclusion

APACHE II score at admission and SA levels with in 24 hrs after admission are independent predictors of mortality for patients with COPD admitted to ICU. The equation derived from these two parameters is useful for predicting outcome of these patients.     

A comparison of the gene expression profiles of CRL-1807 colonocytes exposed to endogenous AAPH-generated peroxides and exogenous peroxides from heated oil

Oxidation of PUFAs in the diet has the potential to be genotoxic and hence carcinogenic. Such carcinogenic processes originate within stem cells of the colon. These cells appear to be predisposed to the carcinogenic process. In colon cells (CRL-1807) exposed to chemical reactions simulating exogenous and endogenous peroxidation reactions, we have observed that undifferentiated cells could mount an effective recombinational repair/TCR response to an endogenous peroxidative DNA damage insult, but not to an external exogenous peroxidative insult as one would encounter from a dietary source. This may suggest that defects in such specific DNA repair may play a role in tumour development in undifferentiated colonocytes exposed to a diet-derived lipid peroxides.