Search

Naturally occurring enzymatic pathways enable highly specific, rapid thiophenic sulfur cleavage occurring at ambient temperature and pressure, which may be harnessed for the desulfurization of petroleum-based fuel. One pathway found in bacteria is a four-step catabolic pathway (the 4S pathway) converting dibenzothiophene (DBT), a common crude oil contaminant, into 2-hydroxybiphenyl (HBP) without disrupting the carbon-carbon bonds. 2'-Hydroxybiphenyl-2-sulfinate desulfinase ( DszB ), the rate-limiting enzyme in the enzyme cascade, is capable of selectively cleaving carbon-sulfur bonds. Accordingly, understanding the molecular mechanisms of DszB activity may enable development of the cascade as industrial biotechnology. Based on crystallographic evidence, we hypothesized that DszB undergoes an active site conformational change associated with the catalytic mechanism. Moreover, we anticipated this conformational change is responsible, in part, for enhancing product inhibition. Rhodococcus erythropolis IGTS8 DszB was recombinantly produced and purified via Escherichia coli BL21 to test these hypotheses. Activity and the resulting conformational change of DszB in the presence of HBP were evaluated. The activity of recombinant DszB was comparable to the natively expressed enzyme and was inhibited via competitive binding of the product, HBP. Using circular dichroism, global changes in DszB conformation were monitored in response to HBP concentration, which indicated that both product and substrate produced similar structural changes. Molecular dynamics (MD) simulations and free energy perturbation with Hamiltonian replica exchange molecular dynamics (FEP/λ-REMD) calculations were used to investigate the molecular-level phenomena underlying the connection between conformation change and kinetic inhibition. In addition to the HBP, MD simulations of DszB bound to common, yet structurally diverse, crude oil contaminants 2',2-biphenol (BIPH), 1,8-naphthosultam (NTAM), 2-biphenyl carboxylic acid (BCA), and 1,8-naphthosultone (NAPO) were performed. Analysis of the simulation trajectories, including root-mean-square fluctuation (RMSF), center of mass (COM) distances, and strength of nonbonded interactions, when compared with FEP/λ-REMD calculations of ligand binding free energy, showed excellent agreement with experimentally determined inhibition constants. Together, the results show that the combination of a molecule's hydrophobicity and nonspecific interactions with nearby functional groups contributes to a competitive inhibition mechanism that locks DszB in a closed conformation and precludes substrate access to the active site.

Background: Poor compliance with chemoprophylaxis is a major contributing factor to the risk of malaria in travelers. Pre-travel chemoprophylaxis may improve compliance by enabling "drug-free holidays." The standard treatment dose of atovaquone/proguanil (250 mg/100 mg, 4 tablets/day for 3 days) provides protection against malaria for at least 4 weeks, and could therefore potentially be used for pre-travel chemoprophylaxis. In this study, we assessed the compliance, tolerability, and acceptability of the 3-day atovaquone/proguanil schedule for malarial chemoprophylaxis., Methods: Two hundred thirty-three participants were recruited from 4 specialized travel medicine clinics in Australia. Adults traveling to malaria-endemic areas with low/medium risk for ≤4 weeks were enrolled and prescribed the 3-day schedule of atovaquone/proguanil, completed at least 1 day before departure. Questionnaires were used to collect data on demographics, travel destination, medication compliance, side effects, and reasons for choosing the 3-day schedule., Results: Overall, 97.7% of participants complied with the 3-day schedule. Although side effects were reported in 43.3% of the participants, these were well tolerated, and mainly occurred during the first and second days. None of the participants developed malaria. The main reasons for choosing the 3-day schedule over standard chemoprophylaxis options were that it was easier to remember (72.1%), required taking fewer tablets (54.0%), and to help scientific research (54.0%)., Conclusions: The 3-day atovaquone/proguanil schedule had an impressively high compliance rate, and was well tolerated and accepted by travelers. Further studies are required to assess the effectiveness of this schedule for chemoprophylaxis in travelers., Clinical Trials Registration: ACTRN12616000640404., (© The Author(s) 2018. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

2'-Hydroxybiphenyl-2-sulfinate (HBPS) desulfinase (DszB) catalyzes the cleavage of the carbon-sulfur bond from HBPS in the final step of microbial 4S pathway desulfurization reactions. DszB is notable for its substrate specificity and exhibits product inhibition, both of which hinder the overall 4S pathway turnover rate. To understand the molecular-level contributions to substrate and inhibitor binding to DszB, we plan to perform molecular dynamic simulations bound to an array of naphthenic molecules and biphenyl analogues of HBPS. However, many of the small molecules we are interested in are not included in standard force field packages, and thus, we must first produce accurate molecular mechanics force fields. Here, we develop and validate CHARMM-compatible force field parameters for the HBPS substrate, the 2-hydroxybiphenyl product, and potential inhibitors including: 2,2'-biphenol, 2-biphenyl carboxylic acid, 1,8-naphthosultam, and 1,8-naphthosultone. The selected molecules represent biphenyl compounds having both a single and double functional group and the planar naphthenic molecule class, all likely present in the oil-rich environment surrounding DszB-producing microorganisms. The Force Field Toolkit (ffTK) in VMD was used to optimize charge, bond distance, angle, and dihedral parameters. Optimized geometries were determined from quantum mechanical calculations. Molecular simulations of the molecules in explicit and implicit water solutions were conducted to assess the abilities of optimized parameters to recapitulate optimized geometries. Calculated infrared (IR) spectra were obtained and compared with experimental IR spectra for validation of the optimized MM parameters., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Background: There are few reports of pediatric studies of atrial fibrillation (AF). We sought to describe the clinical characteristics, management strategies, and recurrence rates and to identify predictors of AF recurrence in a contemporary pediatric population., Methods: A retrospective review was performed of patients ≤ 18 years with lone AF who were seen at 4 pediatric institutions from 1996-2011. Patients with AF in the setting of thyroid disease, ventricular pre-excitation, coexisting congenital heart disease, or a history of cardiac surgery were excluded. Demographics, clinical presentation, investigations, treatment, and follow-up were analyzed., Results: Forty-two patients were diagnosed with a first episode of lone AF, and 4 of these cases were later classified as persistent AF. Thirty-one (74%) were male patients, median age was 15.3 years, and median (interquartile range [IQR]) duration of AF episode was 12 (IQR, 7-24) hours. AF recurred in 39% (15 of 38) of patients. The Kaplan-Meier median time to estimated recurrence was 19 months. By univariate analysis, initial AF episode duration was associated with a higher risk of recurrence (hazard ratio [HR], 1.01; 95% confidence interval [CI], 1-1.02; P = 0.034). Sex, age, family history, size of the left atrium, and history of cardioversion were not associated with recurrence. Recurrence with another supraventricular tachyarrhythmia (SVT) was observed in 6 of 38 (16%) patients, and 12 patients underwent electrophysiology (EP) study, with 6 patients receiving ablation., Conclusions: Our reported rate of recurrence of 39% is important when counseling pediatric patients and their parents on the expected course and treatment goals., (Copyright © 2013 Canadian Cardiovascular Society. Published by Elsevier Inc. All rights reserved.)

Study Objective: To understand adolescent girls' use of "I don't know" within the context of discussing information related to sexual attitudes and behaviors., Design: Qualitative analysis of seven focus groups with adolescent girls., Setting: Urban primary care clinic., Participants: Adolescent girls (n = 23) with a mean age of 16.4 years (range 14 to 18 years). Fifty-two percent were African-American, 26% were Caucasian, 17% were Hispanic/Latino, and 5% were Asian., Results: The use of "I don't know" served three functions: (1) place holder, (2) lack of commitment to an opinion; and (3) reduction of commitment to an opinion or belief., Conclusions: These results suggest that girls use "I don't know" when asked about their thoughts and opinions related to sexual health, and that its use can have varying implications. Discriminating the intent may help providers respond appropriately. Responses should include providing a safe and nonjudgmental environment in which girls can express their feelings and opinions regarding their sexual health.

Recently, a number of "quick tests" became available for use in on-site drug testing. These tests offer advantages in simplicity, ease of performance, and rapid access to test results. However, there is a paucity of data on the validity of these tests for the detection of drugs of abuse. This report describes a validity study of the EZ-SCREEN cannabinoid test for the detection of cannabinoids in urine. Three healthy, male volunteers with a history of marijuana use participated in the study. Each subject smoked 1, 2, or 4 marijuana cigarettes (2.6% THC) on each test day. Urine samples were collected and incorporated into a specimen set consisting of 178 clinical urine samples, 72 urine samples containing known amounts of drug, and 50 drug-free urine samples. The specimen set was randomized and analyzed under blind conditions by the EZ-SCREEN test and by GC/MS for 11-nor-9-carboxy-delta 9-tetrahydrocannabinol (THCCOOH). Results were interpreted independently by three readers. Concordance analysis was performed by comparison of results of the EZ-SCREEN test with GC/MS. The EZ-SCREEN test was highly sensitive and produced positive results at a standard THCCOOH concentration of 5 ng/mL. While showing high sensitivity to THCCOOH, the assay demonstrated low cross-reactivity with delta 9-tetrahydrocannabinol (THC) and other cannabinoids. No false-positive results were recorded with 50 drug-free urine samples, but one reader recorded eight undecided results. Overall agreement between the three readers for the EZ-Screen results was approximately 80%. Delayed readings and photocopy readings tended to be less accurate than readings obtained at 3 min.(ABSTRACT TRUNCATED AT 250 WORDS)

To assess the effects of various clinical factors in determining the cost and length of stay in patients undergoing electrophysiologic testing for cardiac arrhythmias, the hospital cost and length of stay data were reviewed in 222 consecutive inpatients who underwent electrophysiologic testing from January 1 to December 31, 1984. Admissions were classified as: primarily for treatment of arrhythmias (171 patients); primarily for treatment of arrhythmias but with serious concurrent illnesses that prolonged hospitalization (43 patients); or primarily for nonarrhythmic problems with electrophysiologic study an incidental part of hospitalization (8 patients). Based on allowable length of stay for the applicable DRGs, actual hospitalizations exceeded Medicare allowable length of stay by 50 to 500%. Retrospective review of hospital charts indicated that 3 clinical factors serve as effective markers in determining length of stay: need for amiodarone, induction of sustained ventricular tachycardia (VT) or ventricular fibrillation (VF), and presence of serious other medical problems that require stabilization before electrophysiologic testing. Our data indicate that 3 classes of patients can be identified: I. DRG A (45%)--those who did not have sustained VT or VF induced, did not require amiodarone and had no serious concurrent illnesses. The mean length of stay was 7.1 days. II. DRG B patients (21%)--those who had sustained VT or VF induced, but did not require amiodarone and had no serious concurrent illnesses. The mean length of stay was 13.7 days. III. DRG C patients (34%)--those who either had a serious concurrent illness or required amiodarone. The mean length of stay was 19.7 days. This classification schema might allow a more appropriate system for determining reimbursement.