The catalytic residues of Tn3 resolvase

To characterize the residues that participate in the catalysis of DNA cleavage and rejoining by the site-specific recombinase Tn 3 resolvase, we mutated conserved polar or charged residues in the catalytic domain of an activated resolvase variant. We analysed the effects of mutations at 14 residues on proficiency in binding to the recombination site (‘site I’), formation of a synaptic complex between two site Is, DNA cleavage and recombination. Mutations of Y6, R8, S10, D36, R68 and R71 resulted in greatly reduced cleavage and recombination activity, suggesting crucial roles of these six residues in catalysis, whereas mutations of the other residues had less dramatic effects. No mutations strongly inhibited binding of resolvase to site I, but several caused conspicuous changes in the yield or stability of the synapse of two site Is observed by non-denaturing gel electrophoresis. The involvement of some residues in both synapsis and catalysis suggests that they contribute to a regulatory mechanism, in which engagement of catalytic residues with the substrate is coupled to correct assembly of the synapse

Multipart DNA Assembly Using Site-Specific Recombinases from the Large Serine Integrase Family.

Assembling multiple DNA fragments into functional plasmids is an important and often rate-limiting step in engineering new functions in living systems. Bacteriophage integrases are enzymes that carry out efficient recombination reactions between short, defined DNA sequences known as att sites. These DNA splicing reactions can be used to assemble large numbers of DNA fragments into a functional circular plasmid in a method termed serine integrase recombinational assembly (SIRA). The resulting DNA assemblies can easily be modified by further recombination reactions catalyzed by the same integrase in the presence of its recombination directionality factor (RDF). Here we present a set of protocols for the overexpression and purification of bacteriophage ϕC31 and Bxb1 integrase and RDF proteins, their use in DNA assembly reactions, and subsequent modification of the resulting DNA assemblies

The glyoxal budget and its contribution to organic aerosol for Los Angeles, California, during CalNex 2010

Recent laboratory and field studies have indicated that glyoxal is a potentially large contributor to secondary organic aerosol mass. We present in situ glyoxal measurements acquired with a recently developed, high sensitivity spectroscopic instrument during the CalNex 2010 field campaign in Pasadena, California. We use three methods to quantify the production and loss of glyoxal in Los Angeles and its contribution to organic aerosol. First, we calculate the difference between steady state sources and sinks of glyoxal at the Pasadena site, assuming that the remainder is available for aerosol uptake. Second, we use the Master Chemical Mechanism to construct a two-dimensional model for gas-phase glyoxal chemistry in Los Angeles, assuming that the difference between the modeled and measured glyoxal concentration is available for aerosol uptake. Third, we examine the nighttime loss of glyoxal in the absence of its photochemical sources and sinks. Using these methods we constrain the glyoxal loss to aerosol to be 0-5 × 10-5 s-1 during clear days and (1 ± 0.3) × 10-5 s-1 at night. Between 07:00-15:00 local time, the diurnally averaged secondary organic aerosol mass increases from 3.2 μg m-3 to a maximum of 8.8 μg m -3. The constraints on the glyoxal budget from this analysis indicate that it contributes 0-0.2 μg m-3 or 0-4% of the secondary organic aerosol mass. Copyright 2011 by the American Geophysical Union

Formalization of Transform Methods using HOL Light

Transform methods, like Laplace and Fourier, are frequently used for analyzing the dynamical behaviour of engineering and physical systems, based on their transfer function, and frequency response or the solutions of their corresponding differential equations. In this paper, we present an ongoing project, which focuses on the higher-order logic formalization of transform methods using HOL Light theorem prover. In particular, we present the motivation of the formalization, which is followed by the related work. Next, we present the task completed so far while highlighting some of the challenges faced during the formalization. Finally, we present a roadmap to achieve our objectives, the current status and the future goals for this project.Comment: 15 Pages, CICM 201

Atmospheric emissions from the deepwater Horizon spill constrain air-water partitioning, hydrocarbon fate, and leak rate

The fate of deepwater releases of gas and oil mixtures is initially determined by solubility and volatility of individual hydrocarbon species; these attributes determine partitioning between air and water. Quantifying this partitioning is necessary to constrain simulations of gas and oil transport, to predict marine bioavailability of different fractions of the gas-oil mixture, and to develop a comprehensive picture of the fate of leaked hydrocarbons in the marine environment. Analysis of airborne atmospheric data shows massive amounts (∼258,000 kg/day) of hydrocarbons evaporating promptly from the Deepwater Horizon spill; these data collected during two research flights constrain air-water partitioning, thus bioavailability and fate, of the leaked fluid. This analysis quantifies the fraction of surfacing hydrocarbons that dissolves in the water column (∼33% by mass), the fraction that does not dissolve, and the fraction that evaporates promptly after surfacing (∼14% by mass). We do not quantify the leaked fraction lacking a surface expression; therefore, calculation of atmospheric mass fluxes provides a lower limit to the total hydrocarbon leak rate of 32,600 to 47,700 barrels of fluid per day, depending on reservoir fluid composition information. This study demonstrates a new approach for rapid-response airborne assessment of future oil spills. Copyright 2011 by the American Geophysical Union

Bridging Alone: Religious Conservatism, Marital Homogamy, and Voluntary Association Membership

This study characterizes social insularity of religiously conservative American married couples by examining patterns of voluntary associationmembership. Constructing a dataset of 3938 marital dyads from the second wave of the National Survey of Families and Households, the author investigates whether conservative religious homogamy encourages membership in religious voluntary groups and discourages membership in secular voluntary groups. Results indicate that couples’ shared affiliation with conservative denominations, paired with beliefs in biblical authority and inerrancy, increases the likelihood of religious group membership for husbands and wives and reduces the likelihood of secular group membership for wives, but not for husbands. The social insularity of conservative religious groups appears to be reinforced by homogamy—particularly by wives who share faith with husbands

Generalized Theorems for Nonlinear State Space Reconstruction

Takens' theorem (1981) shows how lagged variables of a single time series can be used as proxy variables to reconstruct an attractor for an underlying dynamic process. State space reconstruction (SSR) from single time series has been a powerful approach for the analysis of the complex, non-linear systems that appear ubiquitous in the natural and human world. The main shortcoming of these methods is the phenomenological nature of attractor reconstructions. Moreover, applied studies show that these single time series reconstructions can often be improved ad hoc by including multiple dynamically coupled time series in the reconstructions, to provide a more mechanistic model. Here we provide three analytical proofs that add to the growing literature to generalize Takens' work and that demonstrate how multiple time series can be used in attractor reconstructions. These expanded results (Takens' theorem is a special case) apply to a wide variety of natural systems having parallel time series observations for variables believed to be related to the same dynamic manifold. The potential information leverage provided by multiple embeddings created from different combinations of variables (and their lags) can pave the way for new applied techniques to exploit the time-limited, but parallel observations of natural systems, such as coupled ecological systems, geophysical systems, and financial systems. This paper aims to justify and help open this potential growth area for SSR applications in the natural sciences

African-American crack abusers and drug treatment initiation: barriers and effects of a pretreatment intervention

BACKGROUND: Individual and sociocultural factors may pose significant barriers for drug abusers seeking treatment, particularly for African-American crack cocaine abusers. However, there is evidence that pretreatment interventions may reduce treatment initiation barriers. This study examined the effects of a pretreatment intervention designed to enhance treatment motivation, decrease crack use, and prepare crack abusers for treatment entry. METHODS: Using street outreach, 443 African-American crack users were recruited in North Carolina and randomly assigned to either the pretreatment intervention or control group. RESULTS: At 3-month follow-up, both groups significantly reduced their crack use but the intervention group participants were more likely to have initiated treatment. CONCLUSION: The intervention helped motivate change but structural barriers to treatment remained keeping actual admissions low. Policy makers may be interested in these pretreatment sites as an alternative to treatment for short term outcomes

PICH promotes sister chromatid disjunction and co-operates with topoisomerase II in mitosis

PICH is a SNF2 family DNA translocase that binds to ultra-fine DNA bridges (UFBs) in mitosis. Numerous roles for PICH have been proposed from protein depletion experiments, but a consensus has failed to emerge. Here, we report that deletion of PICH in avian cells causes chromosome structural abnormalities, and hypersensitivity to an inhibitor of Topoisomerase II (Topo II), ICRF-193. ICRF-193-treated PICH-/- cells undergo sister chromatid non-disjunction in anaphase, and frequently abort cytokinesis. PICH co-localises with Topo IIα on UFBs and at the ribosomal DNA locus, and the timely resolution of both structures depends on the ATPase activity of PICH. Purified PICH protein strongly stimulates the catalytic activity of Topo II in vitro. Consistent with this, a human PICH-/- cell line exhibits chromosome instability and chromosome condensation and decatenation defects similar to those of ICRF-193-treated cells. We propose that PICH and Topo II cooperate to prevent chromosome missegregation events in mitosis