Equilibrium and kinetic parameters of the sequence-specific interaction of Escherichia coli RNA polymerase with nontemplate strand oligodeoxyribonucleotides

The specific recognition by Escherichia coli RNA polymerase of single-stranded oligodeoxyribonucleotides (oligos) with the sequence of the -10 promoter region on the nontemplate strand has been studied. Binding was monitored by observing the increase in fluorescence of 2-aminopurine residues incorporated in the oligos. The effects of salt on the rates of formation and dissociation of RNA polymerase.oligo complexes are relatively small, from which we conclude that electrostatic interactions contribute minimally to the favorable binding free energy. From the convex temperature dependence of ln Ka (Ka is the equilibrium association constant), we infer that a large apparent negative heat capacity, of 1-2 kcal M-1 K-1, accompanies complex formation, which is interpreted as due to a conformational change in RNA polymerase. Contrary to expectation, the forward rate constant for binding of oligos is more than 10-fold smaller than that for open complex formation at strong promoters. This suggests that in comparison to an oligo, promoter DNA may be better able to accelerate this required conformational change in the RNA polymerase. Oligo binding was shown to compete with the interaction between RNA polymerase and promoters, indicating that the two bind to overlapping sites on the RNA polymerase     

Myosin conformational states determined by single fluorophore polarization

Muscle contraction is powered by the interaction of the molecular motor myosin with actin. With new techniques for single molecule manipulation and fluorescence detection, it is now possible to correlate, within the same molecule and in real time, conformational states and mechanical function of myosin. A spot-confocal microscope, capable of detecting single fluorophore polarization, was developed to measure orientational states in the smooth muscle myosin light chain domain during the process of motion generation. Fluorescently labeled turkey gizzard smooth muscle myosin was prepared by removal of endogenous regulatory light chain and re-addition of the light chain labeled at cysteine-108 with the 6-isomer of iodoacetamidotetramethylrhodamine (6-IATR). Single myosin molecule fluorescence polarization data, obtained in a motility assay, provide direct evidence that the myosin light chain domain adopts at least two orientational states during the cyclic interaction of myosin with actin, a randomly disordered state, most likely associated with myosin whereas weakly bound to actin, and an ordered state in which the light chain domain adopts a finite angular orientation whereas strongly bound after the powerstroke.     

Subjective symptoms and sleep apnea among persons referred to a sleep center. A questionnaire survey

The aim of this study was to evaluate the relation between symptoms and clinical characteristics and the frequency of increased apnoea and hypopnoea activity. The study population consisted of 135 persons referred to a sleep centre. One hundred and fifteen patients (85%) completed a questionnaire and were subsequently monitored during one night's sleep. Fifty-two persons (45%) had an Apnoea Hypopnoea Index (AHI = the mean number of apnoea and hypopnoea episodes per hour of sleep) > or = 10. Forty-one (36%) had an Apnoea Index (AI = the mean number of apnoea per hour of sleep) > or = 10. Symptoms and clinical characteristics had no significant correlation to increased apnoea activity; however, there was a significant relationship between self-reported frequent breathing pauses during sleep and increased AHI and AI. Symptoms and clinical characteristics are not capable of identifying persons with increased apnoea activity.     

Potent and selective bicyclic lactam inhibitors of thrombin: Part 2: P1 modifications

The synthesis and antithrombotic activity of a series of nonpeptide bicyclic thrombin inhibitors is described. We have explored the SAR with modifications to the P1 site. The introduction of arginine mimetics at the P1 site led to potent and selective thrombin inhibitors.     

Interrelationships among disablement concepts

BACKGROUND: Understanding interrelationships among disablement concepts is critical to the design of future disability treatment and prevention interventions. METHODS: This study uses cross-sectional data to examine the relationships among physiologic impairments, functional limitations, and disability in a moderately disabled sample of 207 community-dwelling older adults. RESULTS: As hypothesized, the data revealed statistically significant curvilinear relationships of upper and lower extremity strength and balance with mobility in this older sample. Multivariate analyses further clarified the hypothesized causal mechanism among the disablement concepts by demonstrating that most of the association of muscle strength and balance with disability was through the intermediary role of mobility limitations. CONCLUSIONS: The findings from this study highlight the value of clinical trials that focus on prevention or treatment of mobility limitations as a means of preventing disability; our findings underscore the need for future research that examines the effects of other variables believed to influence disablement in late life.     

Prenatal ethanol exposure decreases GAP-43 phosphorylation and protein kinase C activity in the hippocampus of adult rat offspring

Consumption of moderate quantities of ethanol during pregnancy produces deficits in long-term potentiation in the hippocampal formation of adult offspring. Protein kinase C (PKC)-mediated phosphorylation of the presynaptic protein GAP-43 is critical for the induction of long-term potentiation. We tested the hypothesis that this system is affected in fetal alcohol-exposed (FAE) rats by measuring GAP-43 phosphorylation and PKC activity in the hippocampus of adult offspring of rat dams that had consumed one of three diets throughout gestation: (a) a 5% ethanol liquid diet, which produced a maternal blood ethanol concentration of 83 mg/dl (FAE); (b) an isocalorically equivalent 0% ethanol diet (pair-fed); or (c) lab chow ad libitum. Western blot analysis using specific antibodies to PKC-phosphorylated GAP-43 revealed that FAE rats had an approximately 50% reduction in the proportion of phosphorylated GAP-43. Similarly, we found that PKC-mediated incorporation of 32P into GAP-43 was reduced by 85% in hippocampal slices from FAE rats compared with both control groups. FAE animals also showed a 50% reduction in total hippocampal PKC activity, whereas the levels of six major PKC isozymes did not change in any of the diet groups. These results suggest that GAP-43 phosphorylation deficits in rats prenatally exposed to moderate levels of ethanol are not due to alterations in the expression of either the enzyme or substrate protein, but rather to a defect in kinase activation.     

Clinical impact of blood cultures taken in the emergency department

OBJECTIVES: (1) To establish the proportion of blood cultures that yield a positive microbiological culture. (2) To determine what proportion of blood cultures taken in the emergency department impact on patient management. (3) To develop guidelines for the appropriate ordering of blood cultures from patients in the emergency department (ED). METHODS: Retrospective review of all blood cultures taken in the ED of a metropolitan teaching hospital between 26 September 1995 and 30 June 1996. RESULTS: One thousand and sixty two blood cultures were taken from patients in the ED during the study period. Ninety two (9%) returned a positive microbiological culture. Of these, 52 (5%) were "true" positives and 18 (1.6%) resulted in changes in management. CONCLUSIONS: Blood cultures taken in the ED rarely yield positive cultures. Only 1.6% of blood cultures taken in the ED impact on management of patients. Simple strategies could reduce the number of blood cultures ordered with little prospect of patient compromise.     

The use of the BiPAP biphasic positive airway pressure system in acute spinal cord injury

Transurethral electrovaporization of the prostate (TVP) is a new minimally invasive procedure for treatment of enlargement of prostate. Between April 1996 and September 1997, TVP was carried out in 109 cases with symptoms of lower urinary tract obstruction. A Stortz spike electrode and a Stortz vapor cutting electrode were used for vaporization electrodes. Efficacy parameters evaluated included International Prostate Symptom Score, peak uroflow and postvoid residual volume. By September 1997, 32 cases (spike electrode) and 33 cases (vapor cutting electrode) could be followed up and evaluated. They have shown both subjective and objective voiding improvement. The improvements in subjective symptom scores and objective voiding parameters were not significantly different between the two electrode groups. Early complications included urinary retention, intraoperative burn, stress incontinence, blood transfusion and postoperative hemorrhage. Late complications included urethral stricture, bladder neck contracture and bladder stone. TVP was found to have several advantages, particularly minimal bleeding and the low incidence of postoperative morbidity. The technique is simple and symptoms improve at an early stage following surgery. This study demonstrates that TVP is a safe and effective modality for treatment of BPH. However, long-term studies with larger numbers of patients are needed.