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To perform double-stranded DNA passage, type II topoisomerases generate a covalent enzyme-cleaved DNA complex (i.e. cleavage complex). Although this complex is a requisite enzyme intermediate, it is also intrinsically dangerous to genomic stability. Consequently, cleavage complexes are the targets for several clinically relevant anticancer and antibacterial drugs. Human topoisomerase IIα and IIβ and bacterial gyrase maintain higher levels of cleavage complexes with negatively supercoiled over positively supercoiled DNA substrates. Conversely, bacterial topoisomerase IV is less able to distinguish DNA supercoil handedness. Despite the importance of supercoil geometry to the activities of type II topoisomerases, the basis for supercoil handedness recognition during DNA cleavage has not been characterized. Based on the results of benchtop and rapid-quench flow kinetics experiments, the forward rate of cleavage is the determining factor of how topoisomerase IIα/IIβ, gyrase and topoisomerase IV distinguish supercoil handedness in the absence or presence of anticancer/antibacterial drugs. In the presence of drugs, this ability can be enhanced by the formation of more stable cleavage complexes with negatively supercoiled DNA. Finally, rates of enzyme-mediated DNA ligation do not contribute to the recognition of DNA supercoil geometry during cleavage. Our results provide greater insight into how type II topoisomerases recognize their DNA substrates.

Cytochrome P450 2D6 (CYP2D6) is a major drug metabolizing enzyme with a buried active site. Channels leading to the active site from various enzyme surfaces are believed to facilitate ligand egress and access to the active site. The present study used molecular dynamics (MD) and in vitro studies with CYP2D6*1 and a Trp75 to Ala mutant to examine channel gating in CYP2D6 by Trp75. MD simulations measured energy landscapes of Trp75 conformations and simulated substrate passage within channel 2b using bufuralol as a model substrate. Trp75 alternated between multiple stable states that supported substrate transport along channel 2b with low energy barriers between states (~-1 kcal/mol). Trp75 conformations were stabilized primarily by hydrogen bonding between Trp75 and Glu222, Asn226, Ala225, or Gln72. Energy barriers were low between Trp75 conformations allowing Trp75 to easily move between various conformations over time and to function in both binding to and moving substrates in the 2b channel of CYP2D6. Michaelis-Menten kinetic studies completed with purified enzyme in a reconstituted system showed overall reduced enzyme efficiency for metabolism of bufuralol and dextromethorphan by the Trp75Ala mutant compared to CYP2D6*1. In stopped-flow measurements, koff for dextromethorphan was decreased in the absence of Trp75. Our results support a role for Trp75 in substrate shuttling to the active site of CYP2D6. Significance Statement Using combined molecular dynamics and in vitro assays, this study shows for the first time a role for Trp75 as a channel entrance gating residue in the mechanism of substrate binding/unbinding in CYP2D6. Energy landscapes derived from molecular dynamics were used to quantitate the strength of gating and kinetics assays showed the impact on enzyme efficiency and koff of a Trp75Ala mutation.

Performance measurement provides an evidence-based means to inform development of interventions to improve the quality of care for people who use opioids. We aimed to develop and assess the predictive validity of health system performance measures for opioid use disorder (OUD) in British Columbia (BC), Canada.Performance measures were generated using retrospective population-level administrative datasets (both provincial and regional) and publicly-reported retrospective data according to four domains (care engagement, clinical guideline compliance, integration, and healthcare utilization). The adjusted odds ratio was estimated via generalized linear mixed models to determine predictive validity for all-cause hospitalization or mortality within 6 months of measurement.A total of 102 performance measures were constructed. We identified 55,470 diagnosed PWOUD, and 39,456 ever engaged in opioid agonist treatment (OAT). We found divergent rates of treatment for concurrent conditions (7.4% for alcohol use disorder to 80.1% for HIV/AIDS), low levels of linkage to OAT and other outpatient care following acute care, and increasing levels of service provision, including increases in OAT prescribers and pharmacies, naloxone kit distribution and overdose prevention site visitation. Our analyses on the predictive validity measures largely supported a priori hypotheses on the direction of effect on the outcome.We identified a range of priorities to improve the quality of care for PWOUD, with critical gaps in linkage to care through acute care settings and long-term engagement in OAT. The proposed measures can be derived for geographic and clinical subgroups and updated over time, providing a basis to monitor and evaluate efforts to address the public health burden of OUD.

It has been recognized for >50 years that cytochrome b5 (b5) stimulates some cytochrome P450 (P450)–catalyzed oxidations, but the basis of this function is still not understood well. The strongest stimulation of catalytic activity by b5 is in the P450 17A1 lyase reaction, an essential step in androgen synthesis from 21-carbon (C21) steroids, making this an excellent model system to interrogate b5 function. One of the issues in studying b5–P450 interactions has been the limited solution assay methods. We constructed a fluorescently labeled variant of human b5 that can be used in titrations. The labeled b5 bound to WT P450 17A1 with a Kd of 2.5 nM and rapid kinetics, on the order of 1 s−1. Only weak binding was observed with the clinical P450 17A1 variants E305G, R347H, and R358Q; these mutants are deficient in lyase activity, which has been hypothesized to be due to attenuated b5 binding. Kd values were not affected by the presence of P450 17A1 substrates. A peptide containing the P450 17A1 Arg-347/Arg-358 region attenuated Alexa 488-T70C-b5 fluorescence at higher concentrations. The addition of NADPH–P450 reductase (POR) to an Alexa 488-T70C-b5:P450 17A1 complex resulted in a concentration-dependent partial restoration of b5 fluorescence, indicative of a ternary P450:b5:POR complex, which was also supported by gel filtration experiments. Overall, these results are interpreted in the context of a dynamic and tight P450 17A1:b5 complex that also binds POR to form a catalytically competent ternary complex, and variants that disrupt this interaction have low catalytic activity.

Cytochrome P450 (P450) 3A4 is the enzyme most involved in the metabolism of drugs and can also oxidize numerous steroids. This enzyme is also involved in one-half of pharmacokinetic drug–drug interactions, but details of the exact mechanisms of P450 3A4 inhibition are still unclear in many cases. Ketoconazole, clotrimazole, ritonavir, indinavir, and itraconazole are strong inhibitors; analysis of the kinetics of reversal of inhibition with the model substrate 7-benzoyl quinoline showed lag phases in several cases, consistent with multiple structures of P450 3A4 inhibitor complexes. Lags in the onset of inhibition were observed when inhibitors were added to P450 3A4 in 7-benzoyl quinoline O-debenzylation reactions, and similar patterns were observed for inhibition of testosterone 6β-hydroxylation by ritonavir and indinavir. Upon mixing with inhibitors, P450 3A4 showed rapid binding as judged by a spectral shift with at least partial high-spin iron character, followed by a slower conversion to a low-spin iron–nitrogen complex. The changes were best described by two intermediate complexes, one being a partial high-spin form and the second another intermediate, with half-lives of seconds. The kinetics could be modeled in a system involving initial loose binding of inhibitor, followed by a slow step leading to a tighter complex on a multisecond time scale. Although some more complex possibilities cannot be dismissed, these results describe a system in which conformationally distinct forms of P450 3A4 bind inhibitors rapidly and two distinct P450–inhibitor complexes exist en route to the final enzyme–inhibitor complex with full inhibitory activity.

Studies on the development of critical thinking skills with specific curriculum materials and instructional methods are few and have been highly theoretical and far removed from practical concerns and applications. This paper contributes to the existing literature on critical thinking pedagogy by providing examples of written assignments that are designed using the 21st century Bloom’s taxonomy model to contribute to critical thinking development. The step-wise development of critical thinking skills approach is provided. The paper argues that both focus on real problems and issues and provision of clear unambiguous instructions are critical fundamentals of critical thinking skills development. The paper also illustrates how to develop critical thinking assignments by providing five examples of course assignments within business education.

We have synthesized polypyrrole nanoparticles using three different oxidizing agents (hydrogen peroxide, chloroauric acid and ferric chloride) and shown that films assembled from these nanoparticles have significantly different drug release profiles. When ferric chloride is used as the oxidizing agent, it is possible to release drugs at voltages as low as -0.05 V, almost an order of magnitude lower than typically used voltages. These ultra-low voltage responsive nanoparticles widen the window of operation of conducting polymers and enable delivery of redox active drugs. As an example, we have shown pulsed release of the chemotherapeutic methotrexate at voltages as low as -0.075 V, demonstrating the potential application of these nanoparticles in cancer treatment. We have also verified the anti-tumor efficacy of the released drug using PC12 cell cultures.

Cytochrome P450 (P450) 17A1 catalyzes the 17α-hydroxylation of progesterone and pregnenolone as well as the subsequent lyase cleavage of both products to generate androgens. However, the selective inhibition of the lyase reactions, particularly with 17α-hydroxy pregnenolone, remains a challenge for the treatment of prostate cancer. Here, we considered the mechanisms of inhibition of drugs that have been developed to inhibit P450 17A1, including ketoconazole, seviteronel, orteronel, and abiraterone, the only approved inhibitor used for prostate cancer therapy, as well as clotrimazole, known to inhibit P450 17A1. All five compounds bound to P450 17A1 in a multistep process, as observed spectrally, over a period of 10 to 30 s. However, no lags were observed for the onset of inhibition in rapid-quench experiments with any of these five compounds. Furthermore, the addition of substrate to inhibitor–P450 17A1 complexes led to an immediate formation of product, without a lag that could be attributed to conformational changes. Although abiraterone has been previously described as showing slow-onset inhibition (t1/2 = 30 min), we observed rapid and strong inhibition. These results are in contrast to inhibitors of P450 3A4, an enzyme with a larger active site in which complete inhibition is not observed with ketoconazole and clotrimazole until the changes are completed. Overall, our results indicate that both P450 17A1 reactions—17α-hydroxylation and lyase activity—are inhibited by the initial binding of any of these inhibitors, even though subsequent conformational changes occur.

Purpose – The purpose of this study/paper is to highlight emerging themes in CRE capabilities and organizational models. The authors explain a Lean Six Sigma-based staffing optimization and integration methodology which ensures organizational design is aligned with enterprise requirements. The ultimate objective is a CRE organizational model that sets resourcing requirements for years ahead while yielding innovative new savings. These priorities set an archetypical precedent for emerging organizational models. The emerging organizational blueprint demonstrates a clear move away from the traditional CRE model by optimizing functional service delivery toward a model that drives business integration, strategy development and creative solution deployment. Design/methodology/approach – Throughout numerous assessments of corporate CRE organizations in the past year, the authors have witnessed emerging themes in CRE priorities and the capabilities required for achieving those priorities. The authors have utilized a Lean Six Sigma-based staffing optimization and integration methodology which ensures that organizational design is aligned with enterprise requirements. Findings – Whereas previous models were built to emphasize functional service delivery, the emerging model is structured to enable a core team to focus on developing strategic relationships and delivering strategic solutions. Functional service delivery is managed directly through a strategic partnership with an outsourced service provider. The emerging model suggests a small, centralized core leadership team. Careful governance and communication protocols must be developed to ensure the duplication and redundancy does not become an issue. Originality/value – Lean Six Sigma has evolved into a business improvement system that has taken hold within many high-performing real estate and facility services – a methodology that emerged from mechanized manufacturing might seem ill-adapted to CRE organizational development. Yet, with guided implementation and disciplined use, we have observed that Lean Six Sigma yields high-impact results in service environments and even with non-technical areas like organization design. This is a unique vantage point that combines engineering technicality against cultural aspects of organizational development.

PurposeMost performance measurement systems and scorecards fall short of communicating strategic value because they do not relate the metrics of real estate to the larger objectives of an organization. This paper aims to show that choosing the right business metrics and monitoring them through effective scorecards identifies and amplifies the competitive value that corporate real estate (CRE) creates for its host company.Design/methodology/approachThis paper identifies the common failures in performance measurement systems and presents a six‐step process for effective scorecarding that is formulated solely on strategic business objectives.FindingsThe paper finds that good performance measurement systems create a direct line of sight between CRE goals and stakeholder interests founded on open, strategic dialogue; refinement to a narrow set of metrics; and commitment to consistent process governance.Practical implicationsThe business transparency of a well‐designed performance measurement system helps to prioritize work and clarifies investment and disinvestment decisions.Originality/valueLeading businesses are increasingly refining the quality of their analytics as the source of competitive advantage. CRE executives who adopt the perspective and process described in this paper can elevate real estate as an integral part of the vision and success of their organizations.

Background This phase IIb study explored the antiviral activity and safety of the investigational CC chemokine receptor 5 (CCR5) antagonist aplaviroc (APL) in antiretroviral-naive patients harbouring R5- or R5X4-tropic virus. Methods A total of 191 patients were randomized 2:2:2:1 to one of three APL dosing regimens or to lamivudine (3TC)/zidovudine (ZDV) twice daily (bid), each in combination with lopinavir/ritonavir (LPV/r) 400 mg/100 mg bid. Efficacy, safety and pharmacokinetic parameters were assessed. Results This study was terminated prematurely because of APL-associated idiosyncratic hepatotoxicity. A total of 141 patients initiated treatment early enough to have been able to complete 12 weeks on treatment [modified intent-to-treat (M-ITT) population]; of these, 133 completed the 12-week treatment phase. The proportion of subjects in the M-ITT population with HIV-1 RNA

Recent work by the author has resulted in a computer package which allows the estimation of thermal physical properties of a fluid when very little information is known about that fluid. The minimum required data set for the method is the molecular weight, the density, and temperature of the liquid at the normal boiling point. The method utilizes the application and modification of extended corresponding states. Using a well known reference fluid, or fluids, and the above minimum data set for a target fluid, (the fluid for which properties are desired), thermodynamic and transport properties may be estimated over a wide range of pressure and temperature. Depending on the target fluid and the availability of a suitable reference fluid, the method may use as many as five reference fluids but usually uses three or less. Although the minimum data set for the target fluid is described above, the availability of more data for the target fluid greatly improves the accuracy of the method. Even an additional one or two data points for the target fluid helps and the more data available for the target fluid, the greater the help. The goal of this work was to develop a method and subsequent computer package which would provide thermodynamic and transport properties for a target fluid with a minimal amount of known data for that target fluid, and that the resulting properties for that target fluid be of sufficient accuracy to be useful in engineering calculations. It is believed that this goal has been achieved. Typical accuracy assessments and examples are presented. Since traditionally transport properties have not corresponded well, and since presenting both the thermodynamic and transport property methods would make this paper much to long, only the transport properties part of the package is presented here.

PurposeThe purpose of this paper is to offer a practice guide for where to apply six sigma process improvement methodology to the corporate real estate function.Design/methodology/approachBased on years of direct experience provide practical guidance on how to get started and to overcome resistance that process improvement is not for service organizations like CRE.FindingsWorld‐leading CRE departments are addressing cost pressure and performance expectations using process improvement, and the method of choice is six sigma. A methodology that emerged from mechanized manufacturing might seem ill‐adaptive to CRE and similar service cultures. Yet, with guided implementation and disciplined use, six sigma yields tangible results in service environments.Research limitations/implicationsDocumenting and benchmarking process improvement results across CRE departments; surveying the level of business adoption of six sigma methodology compared to the level of CRE adoption within the same company and how this leads to business misalignmentPractical implicationsWorld class benchmarks will be set in the future by CRE organizations that adopt six sigma process improvement methodology.Originality/valueA provocative discourse furthering the cause six sigma within an industry that famously resistant to change.

Retailers were surveyed to determine their attitudes about energy conservatlon. This paper describes the findings and compares them to results obtained in a previous study of retailers’ energy conservation attitudes which was reported at the Third Annual Conference of the Academy of Marketing Science.

PurposeTo provide a roadmap for corporate real estate (CRE) directors to effect change in the way they deliver services through enhanced relationships with their “customers,” the users of CRE.Design/methodology/approachPrevious studies and discussions of customer relationship management are combined with the authors' extensive experience to make a case for formalized change management processes and analyze specific methods for enhancing the effectiveness of these processes.FindingsRelationship management integrates real estate processes into larger corporate processes, converting real estate opportunities into competitive advantages. The transition from a reactive to a proactive mode of service is aided by adoption of new mind sets and new skills sets, including tools and processes for communicating within the organization, enhanced financial/analytical acumen, and metrics for determining successful outcomes.Research limitations/implicationsEvery CRE department faces unique challenges and opportunities which cannot be fully addressed in a paper designed for broad applicability.Practical implicationsCRE directors will recognize the challenges and opportunities they face and will learn about specific actions and initiatives they can implement to enhance their effectiveness though internal customer relationship management.Originality/valueApplicability of change and customer relationship management processes to the CRE environment is a new area of focus and is mostly uncharted territory. This roadmap delves deeper than previous papers into specific methods for aligning real estate processes with the “voice of the customer” to further corporate objectives.

Corrigendum to “Development of a novel antibody to Calcitonin Gene-Related Peptide for the treatment of osteoarthritis-related pain” [Osteoarthritis Cartilage 22 (2014) 578e585] R.J. Benschop y , E.C. Collins y, R.J. Darling y, B.W. Allan z, D. Leung z, E.M. Conner z, J. Nelson z, B. Gaynor z, J. Xu z, X.-F. Wang z, R.A. Lynch y, B. Li y, D. McCarty y, E.S. Nisenbaum y, J.L. Oskins y, C. Lin y, K.W. Johnson y, M.G. Chambers y

The limitation of available land for wastewater treatment facilities has challenged environmental engineers in searching for efficient and effective treatment systems that will minimize space requirements for waste treatment. To meet these requirements, a fixed film biological process using a DeepBedTM filter for aerobic and anoxic treatment was developed. The innovative biotechnology uses sand as the media to which microorganisms attach and as the filter media for solids retention. The unique characteristics of the biosystem combine long sludge age and short hydraulic detention time and provide small footprints for the bioreactor. The novel technology has been successfully practiced for industrial and municipal wastewater treatment in three (3) continents. This article gives details of a US facility to treat coke plant effluent with high ammonia and phenol content, describes a plant in Taiwan for polishing plastic manufacturing wastewater by combining ozone and biofiltration, illustrates nitrogen removal from a semiconductor factory in Korea, and shows the results of a municipal treatment plant in Britain using BAF to pretreat peak loads from the dairy industry.

A 63-year-old female developed respiratory failure and was admitted to the Intensive Care Unit for non-invasive ventilation, inotropic support and antibiotic therapy. The patient was initially stable but then suddenly deteriorated with acute pulmonary oedema requiring mechanical ventilation. An electrocardiogram showed an acute ST elevation myocardial infarction and the patient subsequently had an urgent coronary angiogram which revealed normal coronary arteries but apical ballooning characteristic of Takotsubo cardiomyopathy. A short review is provided of this relatively newly described heart syndrome which has the potential to present in numbers of intensive care patients. This case emphasises the importance of being aware of uncommon causes of acute ECG changes in the critically ill.

This article describes the phenomenon of morphological instability in solid-solid phase transformations during thin-film interdiffusion, specifically related to the initial stages of precipitation when phase growth occurs along the interface between thin films. The experimental observations that revealed this effect will be presented, and a working hypothesis will be discussed. Experimental observations suggest that the ledge mechanism of growth is present in this system but does not inhibit the formation of interfacial instabilities. It is proposed that morphological stability for solid-phase growth occurring during thin-film interdiffusion can be treated by the inclusion of a solute-source term into the two-dimensional perturbation approach generally used to study unstable growth morphologies. Experimental observations that provide qualitative and semiquantitative support for the solute-source model are also presented.

Se describe una nueva especie mexicana de Aphodiusx A. burgosi Deloya & McCarty, procedente del estado de Moreioe, se comenta eu distribución atfpica dentro del subgénero Coelobrachelus y ee incluye una clave para separar a las siete especies conocidas. la hembra de PanGhrysína parapatríca Deloya & Morón ee descrita por primera vez y se comentan los huéspedes que frecuentan las hembras y loe machos de esta especie. Se incluye una clave para separwr a las seis especies conocidas.

A revised interim formulation for the thermodynamic properties of air has been developed for calculating properties of the vapor and estimating properties for the liquid at temperatures as low as 60 K. The formulation incorporates separate equations for the calculation of bubble-point and dew-point pressures and densities and for the ideal-gas heat capacity. A new fundamental equation of state is given for vapor and liquid states of air based upon available experimental data and predicted values of isochoric heat capacity for the liquid using corresponding states methods. Procedures for predicting Cv are discussed. The fundamental equation for air is explicit in nondimensional Helmholtz energy. The terms of the fundamental equation were selected from a larger set of 75 proposed terms using a least-squares fitting procedure. Representative graphical comparisons of calculated property values to experimental measurements are given. The estimated accuracy of calculated densities is generally ± 0.2% except near the dew and bubble lines. Calculated heat capacities for the liquid must be considered only as estimates until substantiated by experimental measurements.