Kinetic analysis of the coding properties of O6-methylguanine in DNA: the crucial role of the conformation of the phosphodiester bond

Production by N-nitroso compounds of O6-alkylguanine (O6-alkylG) in DNA directs the misincorporation of thymine during DNA replication, leading to G:C to A:T transition mutations, despite the fact that DNA containing O6-alkylG:T base pairs is less stable than that containing O6-alkylG:C pairs. We have examined the kinetics of incorporation by Klenow fragment (KF) of Escherichia coli DNA polymerase I of thymine (T) and of cytosine (C) opposite O6-MeG in the template DNA strand. Both T and C were incorporated opposite O6-MeG much slower than nucleotides forming regular A:T or G:C base pairs. Using various concentrations of dTTP, dCTP, or their phosphorothioate (Sp)-dNTP alpha S analogues, or a mixture of dTTP and dCTP, the progress of incorporation of a single nucleotide in a single catalytic cycle of a preformed KF-DNA complex was measured (pre-steady-state kinetics). The results were consistent with the kinetic scheme (Kuchta, R. D., Benkovic, P., & Benkovic, S. J. (1988) Biochemistry 27, 6716-6725): (1) binding of dNTP to polymerase-DNA; (2) conformational change in polymerase; (3) formation of phosphodiester between the dNTP and the 3'-OH of the primer; (4) conformational change of polymerase; (5) release of pyrophosphate. The results were analyzed mathematically to identify the steps at which the rate constants differ significantly between the incorporation of T and C. The only significant difference was the 5-fold difference in the rates of formation of the phosphodiester bond (for dTTP, kforward = 3.9 s-1 and kback = 1.9 s-1; for dCTP, kforward = 0.7 s-1 and kback = 0.9 s-1). These pre-steady-state progress curves were biphasic with a rapid initial burst followed by an apparently steady-state rise. Deconvolution of these curves gave direct evidence for the importance of the conformational change after polymerization by showing that the curves represented the sum of the rapid accumulation of the product of step 3 followed by the slow conversion of that to the product of step 5 (because of the rapidity of the release of pyrophosphate there was no significant accumulation of the product of step 4). The equilibrium constants for each step suggest that the greatest change in the Gibbs free energy occurs at the conformational change after polymerization and that while the formation of the phosphodiester bond to T is slightly exothermic, that to C is slightly endothermic.(ABSTRACT TRUNCATED AT 400 WORDS)     

Differentiating anxiety and depression in older adults with generalized anxiety disorder.

To examine affect and cognition in differentiating anxiety and depression, 83 older participants with generalized anxiety disorder completed the Cognitive Checklist (CCL) and the Positive and Negative Affect Schedule (PANAS). A 3-factor solution was found for the PANAS: positive affect (PA), anxiety and anger (Negative Affect 1 [NA-1]), and guilt and shame (Negative Affect 2 [NA-2]). A 2-factor structure was noted for the CCL. Correlations with anxiety and depression measures suggested that the CCL Depression (CCL-D) subscale showed stronger correlations with depression, whereas the CCL Anxiety subscale did not uniquely correlate with anxiety. The NA-1 subscale correlated positively with measures of depression and anxiety, whereas the PA subscale showed negative correlations. Hierarchical regression suggested that the CCL-D subscale was a significant predictor of self-reported depression. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

The Estimation of Restrainers

A quantitative method is described for the chromatographic separation and subsequent polarographic estimation of adenine in gelatin. The method, which has been applied to about 200 gelatins, has two important features. It is specific for adenine and enables a visual assessment of other purines and pyrimidines to be mode in the course of the procedure.

Hide gelatins have adenine contents from 10 200 ppm., but the large majority are from SO 50 ppm., ossein gelatin contains from 4–6 ppm., and pigskin between 30 and 50 ppm.     

Sperm phospholipase Czeta from humans and cynomolgus monkeys triggers Ca2+ oscillations,activation and development of mouse oocytes

Fusion with a fertilizing spermatozoon induces the mammalian oocyte to undergo a remarkable series of oscillations in cytosolic Ca(2+), leading to oocyte activation and development of the embryo. The exact molecular mechanism for generating Ca(2+) oscillations has not been established. A sperm-specific zeta isoform of phospholipase C (PLCzeta) has been identified in mice. Mouse PLCzeta triggers Ca(2+) oscillations in mouse oocytes and exhibits properties synonymous with the 'sperm factor' that has been proposed to diffuse into the oocyte after gamete fusion. The present study isolated the PLCzeta homologue from human and cynomolgus monkey testes. Comparison with mouse and monkey PLCzeta protein sequences indicates a shorter X-Y linker region in human PLCzeta and predicts a distinctly different isoelectric point. Microinjection of complementary RNA for both human and cynomolgus monkey PLCzeta elicits Ca(2+) oscillations in mouse oocytes equivalent to those seen during fertilization in mice. Moreover, human PLCzeta elicits mouse egg activation and early embryonic development up to the blastocyst stage, and exhibits greater potency than PLCzeta from monkeys and mice. These results are consistent with the proposal that sperm PLCzeta is the molecular trigger for egg activation during fertilization and that the role and activity of PLCzeta is highly conserved across mammalian species.     

Maximum Likelihood Estimation Using Parallel Computing: An Introduction to MPI

The computational difficulty of econometric problems has increased dramatically in recent years as econometricians examine more complicated models and utilize more sophisticated estimation techniques. Many problems in econometrics are `embarrassingly parallel' and can take advantage of parallel computing to reduce the wall clock time it takes to solve a problem. In this paper I demonstrate a method that can be used to solve a maximum likelihood problem using the MPI message passing library. The econometric problem is a simple multinomial logit model that does not require parallel computing but illustrates many of the problems one would confront when estimating more complicated models.     

Sedimentary records of sewage pollution using faecal markers in contrasting peri-urban shallow lakes

Sewage contamination in shallow lake sediments is of concern because the pathogens, organic matter and nutrients contribute to the deterioration of the water-bodies' health and ecology. Sediment cores from three shallow lakes (Coneries, Church and Clifton Ponds) within Attenborough nature reserve located downstream of sewage treatment works were analysed for TOC, C/N, δ¹³C, δ¹⁵N, bacterial coliforms and faecal sterols. ²¹⁰Pb and ¹³⁷Cs activities were used to date the sediments. Elemental analysis suggests that the source of organic matter was algal and down profile changes in δ¹³C indicate a possible decrease in productivity with time which could be due to improvements in sewage treatment. δ¹⁵N for Coneries Pond are slightly higher than those observed in Church or Clifton and are consistent with a sewage-derived nitrate source which has been diluted by non-sewage sources of N. The similarity in δ¹⁵N values (+ 12‰ to + 10‰) indicates that the three ponds were not entirely hydrologically isolated. Analysis by gas chromatography/mass spectrometry (GC/MS) reveals that Coneries Pond had sterol concentrations in the range 20 to 30 μg/g (dry wt.), whereas, those from Clifton and Church Ponds were lower. The highest concentrations of the human-sourced sewage marker 5β-coprostanol were observed in the top 40 cm of Coneries Pond with values up to 2.2 μg/g. In contrast, Church and Clifton Pond sediments contain only trace amounts throughout. Down-profile comparison of 5β-coprostanol/cholesterol, 5β-coprostanol/(5β-coprostanol + 5α-cholestanol) and 5β-epicoprostanol/coprostanol as well as 5α-cholestanol/cholesterol suggests that Coneries Pond has received appreciable amounts of faecal contamination. Examination of 5β-stigmastanol (marker for herbivorous/ruminant animals) down core concentrations suggests a recent decrease in manure slurry input to Coneries Pond. The greater concentration of β-sitosterol in sediments from Church and Clifton Ponds as compared to Coneries is attributed in part to their greater diversity and extent of aquatic plants and avian faeces.     

Self-linking and self-competence as dimensions of global self-esteem: initial validation of a measure

Three studies were conducted to validate the conceptualization of global self-esteem as consisting of two dimensions: a sense of social worth, or self-liking, and a sense of personal efficacy, or self-competence. In Study 1, confirmatory factor analysis was used to test the a priori structure of the Self-Liking/Self-Competence Scale, a self-report instrument designed to measure the two dimensions. In Study 2, a second structural analysis showed the dimensionality of Rosenberg's (1965) Self-Esteem Scale to parallel the proposed dichotomy. In Study 3, self-liking and self-competence were related to several theoretically linked constructs--depression, self-perceived abilities, and perceived parental approval--with the resulting pattern of correlations supporting their conceptualization as substantively distinct dimensions. The implications of these findings for understanding global self-esteem are discussed.     

Specific sleepiness symptoms are indicators of performance impairment during sleep deprivation

Drivers are not always aware that they are becoming impaired as a result of sleepiness. Using specific symptoms of sleepiness might assist with recognition of drowsiness related impairment and help drivers judge whether they are safe to drive a vehicle, however this has not been evaluated. In this study, 20 healthy volunteer professional drivers completed two randomized sessions in the laboratory – one under 24 h of acute sleep deprivation, and one with alcohol. The Psychomotor Vigilance Task (PVT) and a 30 min simulated driving task (AusEdTM) were performed every 3–4 h in the sleep deprivation session, and at a BAC of 0.00% and 0.05% in the alcohol session, while electroencephalography (EEG) and eye movements were recorded. After each test session, drivers completed the Karolinska Sleepiness Scale (KSS) and the Sleepiness Symptoms Questionnaire (SSQ), which includes eight specific sleepiness and driving performance symptoms. A second baseline session was completed on a separate day by the professional drivers and in an additional 20 non-professional drivers for test–retest reliability. There was moderate test–retest agreement on the SSQ (r = 0.59). Significant correlations were identified between individual sleepiness symptoms and the KSS score (r values 0.50–0.74, p < 0.01 for all symptoms). The frequency of all SSQ items increased during sleep deprivation (χ² values of 28.4–80.2, p < 0.01 for all symptoms) and symptoms were related to increased subjective sleepiness and performance deterioration. The symptoms “struggling to keep your eyes open”, “difficulty maintaining correct speed”, “reactions were slow” and “head dropping down” were most closely related to increased alpha and theta activity on EEG (r values 0.49–0.59, p < 0.001) and “nodding off to sleep” and “struggling to keep your eyes open” were related to slow eye movements (r values 0.67 and 0.64, p < 0.001). Symptoms related to visual disturbance and impaired driving performance were most accurate at detecting severely impaired driving performance (AUC on ROC curve of 0.86–0.91 for detecting change in lateral lane position greater than the change at a BAC of 0.05%). Individual sleepiness symptoms are related to impairment during acute sleep deprivation and might be able to assist drivers in recognizing their own sleepiness and ability to drive safely.     

The fleeting gleam of praise: Cognitive processes underlying behavioral reactions to self-relevant feedback.

We propose that a preference for favorable social feedback (i.e., self-enhancement) requires only that feedback be characterized as favorable or unfavorable but that a preference for self-confirming feedback (i.e., self-verification) is based on a more elaborate set of cognitive operations that requires both the characterization of feedback and a subsequent comparison of that feedback to a representation of self stored in memory. Study 1 set the stage for testing this hypothesis by showing that depriving people of processing resources interfered with their tendency to access their self-conceptions. In Studies 2 and 3, participants who were deprived of resources preferred the favorable, self-enhancing evaluator, whereas control participants displayed a preference for the self-verifying evaluator, even if that evaluator was relatively unfavorable. (PsycINFO Database Record (c) 2010 APA, all rights reserved)