Monomers of human beta 1 beta 1 alcohol dehydrogenase exhibit activity that differs from the dimer

A previously unreported enzymatic activity is described for monomers of the beta 1 beta 1 isoenzyme of human alcohol dehydrogenase that were prepared from dimeric enzyme by freeze-thaw in liquid nitrogen. Whereas the dimeric enzyme has optimal activity at low substrate concentrations (2.5 mM ethanol, 50 microM NAD+; "low Km" activity), the monomer has its highest activity at high substrate concentrations (1.5 M ethanol, 2.5 mM NAD+; "high Km" activity). While the activity of the monomer does not appear to be saturated at 1.5 M ethanol, its maximal activity at this high ethanol concentration exceeds the Vmax of the dimer by about 3-fold. The apparent Km of NAD+ with monomers is 270 microM, and no activity could be detected with nicotinamide mononucleotide as cofactor. During gel filtration the high Km activity elutes at a lower apparent molecular weight position than the dimer. The kinetics of monomer-to-dimer reassociation are consistent with a second-order process with a rate constant of 240 M-1 s-1. The reassociation rate is markedly enhanced by the presence of NAD+. During refolding of beta 1 beta 1 following denaturation in 6 M guanidine hydrochloride, an enzyme species with high Km activity and spectral properties similar to the freeze-thaw monomer is observed, indicating that a catalytically active monomer is an intermediate in the refolding pathway. The enzymatic activity of the monomer implies that the intersubunit contacts of beta 1 beta 1 are not crucial in establishing a catalytically competent enzyme. However, the differences in specific activity and Km between monomer and dimer suggest that dimerization may serve to modulate the catalytic properties.     

Habituation of the orienting response: a gating mechanism subserving selective attention

Possibilities of involution of changes in lesser circulation after closure of experimental aortopulmonary anastomosis were studied. 37 observations at various intervals after closure of anastomosis (several minutes to 13.5 months) in 25 dogs were analyzed. Before closure the anastomosis had functioned for 1-7 months. The results of histological examinations of lungs, pressure measurements in lesser circulation, heart weight, electrocardiographic and spirographic examinations were analyzed. It was found that complete involution of changes in lesser circulation was possible only in first month of existence of anastomosis, in this case with changes of both "early" and "late" types. "Late"-type changes after four months function of anastomosis had both reversible and irreversible character, whereas "early"-type changes became irreversible already after three-month duration of anastomosis. With the "late"-type changes, the operation itself (closure of anastomosis) was accompanied by symptoms of pulmonary vasomotor paresis and heart failure, whereas in the presence of "early"-type changes the operation elicited no morphological or functional changes.     

Radiation- and Photo-Induced Oxidation Pathways of Methionine in Model Peptide Backbone under Anoxic Conditions

Within the reactive oxygen species (ROS) generated by cellular metabolisms, hydroxyl radicals (HO^•) play an important role, being the most aggressive towards biomolecules. The reactions of HO^• with methionine residues (Met) in peptides and proteins have been intensively studied, but some fundamental aspects remain unsolved. In the present study we examined the biomimetic model made of Ac-Met-OMe, as the simplest model peptide backbone, and of HO^• generated by ionizing radiation in aqueous solutions under anoxic conditions. We performed the identification and quantification of transient species by pulse radiolysis and of final products by LC-MS and high-resolution MS/MS after γ-radiolysis. By parallel photochemical experiments, using 3-carboxybenzophenone (CB) triplet with the model peptide, we compared the outcomes in terms of short-lived intermediates and stable product identification. The result is a detailed mechanistic scheme of Met oxidation by HO^•, and by CB triplets allowed for assigning transient species to the pathways of products formation.     

Effect of Tempering and Strain on Decomposition of Metastable Austenite in X210CrW12 Thixo-Cast Steel

Thixoforming of hot rolled X210CrW12tool steel led to the formation of globular austenitic grains (82.4 vol.%) surrounded by eutectic mixture (α-Fe and M₇C₃ carbides). The thixo-cast steel reached compression strength 4.8 GPa at plastic strain 34%. The analysis of pole figures after deformation indicated distinct texturization of microstructure in comparison with undeformed steel. Main texture components for austenite were {101}, 〈010〉, while ferrite did not reveal clearly formed orientation. DSC analysis confirmed that austenitic structure in the X210CrW12 steel was metastable and temperature of decomposition depended on the strain applied at 634 °C for the un-deformed sample and at 599 °C for sample compressed up to 4.8 GPa. Discontinuous transformation of austenite into perlite, that started mainly at grain boundaries and proceeded to the center, was the predominant mechanism responsible for the decomposition of globular grains in thixoformed X210CrW12 steel. The decomposition caused by tempering of supersaturated and severely strained steel led to obtaining characteristic product of transformation of higher hardness in comparison with only tempered sample. In the deformed sample the reaction started on slip bands and twins which revealed high density of defects, promoting precipitation of carbides, followed by local depletion in carbon as a result of α′- Fe formation. In contrast to non-deformed state they covered the area of grains. Two fronts of reaction α-Fe plate +M₃C → mixture of α-Fe and M₇C₃ carbides were also observed.     

Analysis of 59 cases of non-traumatic sudden death in various neurological diseases]

Sudden death is a common complication of myocardial infarction, necrotic pancreatitis and other diseases. Physicians usually neglect the possibility of neurological disorders. 59 cases of sudden death among 314 autopsied cases with neurological diseases were analyzed. The most frequent cause of neurological sudden death was cerebrovascular disease (CVD). It was present in 53 (89.9%) cases. 48 of them had hemorrhagic CVD. 37 of these 48 cases were due to hypothalamic lesions. In 37 cases general autopsy was performed; pathological abnormalities of heart, lungs, stomach etc, were found in 28 of them. In conclusion, the size and location of the lesion correlated with the prognosis of the disease. Concomitant multiple organ damage may deteriorate the lesion. Awareness of sudden death resulting from CVD may elevate the rate of correct diagnosis.     

Dose of hemodialysis and survival: differences by race and sex

CONTEXT: Although blacks receive lower doses of hemodialysis than whites, their survival when receiving dialysis treatment is better than that for whites. Previous studies of the relationship between the dose of dialysis and patient survival have not controlled for differences in patient characteristics. OBJECTIVE: To examine the association of mortality with the dose of hemodialysis for clusters of patients categorized by race and sex. DESIGN: Retrospective analysis of laboratory data and mortality outcomes from 1994, using a national database of hemodialysis patients. PATIENTS: A total of 18144 black and white patients receiving hemodialysis 3 times weekly who either lived the entire year receiving hemodialysis or died. MAIN OUTCOME MEASURES: The fractional reduction of urea in a single dialysis session as the measured hemodialysis dose (urea reduction ratio [URR]) after controlling for race, sex, age, and diabetes mellitus. Mortality was determined by strata of URRs and albumin and creatinine levels. RESULTS: Across all age categories, blacks had lower URRs than whites, and men had lower URRs than women. In an age-adjusted model for evaluating interactions among URRs, race, sex, and diabetes, the association of URR with mortality risk was weak among blacks, particularly black men. After adjustment for age and diabetes, death probability curves were most steep for white women with URR values less than 60%. The death probability curves were least steep for black men. There was no meaningful difference between death probability and albumin or creatinine concentration among the race by sex clusters. CONCLUSION: Using URR, the usual measure of hemodialysis dose, the assumption that the association between dialysis dose and survival is uniform across demographic groups appears incorrect. Comparisons of the quality of dialysis patient care should not rely on URR alone to predict patient survival.     

Specificity of the Speed''s test: arthroscopic technique for evaluating the biceps tendon at the level of the bicipital groove

A historical cohort study was conducted in Misasa town, Tottori prefecture, Japan, where radon spas have been operating for a long time. Misasa town was divided into an elevated radon level area and a control area, with mean indoor radon levels of about 60 and 20 Bq/m3, respectively. In total, 3,083 subjects in the elevated radon level area and 1,248 in the control area, all aged 40 or older on January 1, 1976, were followed up until December 31, 1993, for a mean period of 14 years. The mortality rates from all causes exhibited no difference between the elevated radon level area and the control area for both sexes. No difference was observed in the incidence of all-site cancers (age, period-adjusted rate ratios by Poisson regression, RR = 1.06, 95% confidence interval (CI) 0.79-1.42 for males, RR = 0.90, 95% CI 0.65-1.24 for females), while stomach cancer incidence seemed to decrease for both sexes (RR = 0.70, 95% CI 0.44-1.11 for male, RR = 0.58, 95% CI 0.34-1.00 for female) and lung cancer incidence for males only seemed to increase (RR = 1.65, 95% CI 0.83-3.30 for male, RR = 1.07, 95% CI 0.28-4.14 for female) in the elevated radon level area. Caution is needed in the interpretation of these findings, however, since the individual exposure level was not measured and major confounding factors, such as smoking and diet, could not be controlled in this study.     

Mechanism-based inactivation of human liver cytochrome P450 2A6 by 8-methoxypsoralen

The P450 2A6 catalyzed 7-hydroxylation of coumarin proceeded with a mean Km of 0.40 (+/-0.13) microM and Vmax of 6.34 nmol/nmol P450/min (36-fold variation) in microsomal preparations from a panel of 12 human livers. Substrate depletion was avoided during the kinetic determinations. 8-Methoxypsoralen (8-MOP) is a potent mechanism-based inactivator of human liver P450 2A6 and reconstituted purified recombinant P450 2A6 based on the following evidence: 1) 8-MOP causes time, concentration, and NADPH-dependent loss of P450 2A6 activity that is not reversed by potassium ferricyanide or extensive dialysis, 2) loss of P450 2A6 activity is associated with a loss of spectrally observable P450, 3) addition of nucleophiles or reactive oxygen scavengers to the incubations does not prevent inactivation of P450 2A6, and 4) 8-MOP-dependent P450 2A6 inactivation is inhibited (concentration dependent) by the addition of a competitive inhibitor (pilocarpine). Inactivation is selective for P450 2A6 at low concentrations of 8-MOP (2.5 microM) after short incubation time periods (3 min) and was characterized by a KI of 0.8 and 1.9 microM in a reconstituted and microsomal system, respectively, and a kinact of 1 min-1 and 2 min-1 in a reconstituted and microsomal system, respectively. A substrate depletion partition ratio of 21 was calculated for the inactivation of recombinant P450 2A6. Potency and selectivity suggest that 8-MOP could be a useful tool in vitro for evaluating P450 2A6 activity in various enzyme preparations.     

Racial differences in incidence, outcome, and quality of life for African-Americans on hemodialysis

African-Americans are the fastest growing racial minority with end-stage renal disease (ESRD) in the United States. Although comprising < 15% of the US population, African-Americans comprise approximately 31% of the ESRD population. African-Americans are approximately 7 years younger than their white counterparts with ESRD. Surprisingly, although African-Americans systematically receive less dialysis than whites, their survival with ESRD is greatly improved. This improved survival with ESRD is accompanied by an improved quality of life for African-Americans. In this review, we will examine the reasons why African-Americans have an excessive incidence of selective diseases that culminate in ESRD. Furthermore, we will judge several hypotheses that may account for the improved survival enjoyed by African-Americans with ESRD. We contend that research targeted to clarify the basis for these differences between African-Americans and whites with ESRD will improve the outcomes for both populations.     

Mitochondrial transmission during mating in Saccharomyces cerevisiae is determined by mitochondrial fusion and fission and the intramitochondrial segregation of mitochondrial DNA

To gain insight into the process of mitochondrial transmission in yeast, we directly labeled mitochondrial proteins and mitochondrial DNA (mtDNA) and observed their fate after the fusion of two cells. To this end, mitochondrial proteins in haploid cells of opposite mating type were labeled with different fluorescent dyes and observed by fluorescence microscopy after mating of the cells. Parental mitochondrial protein markers rapidly redistributed and colocalized throughout zygotes, indicating that during mating, parental mitochondria fuse and their protein contents intermix, consistent with results previously obtained with a single parentally derived protein marker. Analysis of the three-dimensional structure and dynamics of mitochondria in living cells with wide-field fluorescence microscopy indicated that mitochondria form a single dynamic network, whose continuity is maintained by a balanced frequency of fission and fusion events. Thus, the complete mixing of mitochondrial proteins can be explained by the formation of one continuous mitochondrial compartment after mating. In marked contrast to the mixing of parental mitochondrial proteins after fusion, mtDNA (labeled with the thymidine analogue 5-bromodeoxyuridine) remained distinctly localized to one half of the zygotic cell. This observation provides a direct explanation for the genetically observed nonrandom patterns of mtDNA transmission. We propose that anchoring of mtDNA within the organelle is linked to an active segregation mechanism that ensures accurate inheritance of mtDNA along with the organelle.