THEORETICAL SOLVATION MODELS:AB INITIO STUDY OF MOLECULAR AGGREGATION

The results of a comparative ab initio study of solvation effects in a process of hydrogen bonddriven dimerisation of two small organic molecules - formamide and a primary amide of the α-hydroxyaceticacid are here presented. Differences between various dielectric continuum models (i.e. Onsager, PCM, IPCMand SCIPCM), their performance and range of applicability are reported and discussed.Pozna

Protein Structure Analysis

We describe a series of engaging exercises in which students emulate the process that researchers use to efficiently develop new pharmaceutical drugs, that of rational drug design. The activities are taken from a three- to four-hour workshop regularly conducted with first-year college students and presented here to take place over three to four class periods. Although targeted at college students, these activities may be appropriate at the high school level as well, particularly in an AP Biology course. The exercises introduce students to the topics of bioinformatics and computer modeling, in the context of rational drug design, using free online resources such as databases and computer programs. Through the process of learning about computational drug design and drug optimization, students also learn content such as elements of protein structure and protein–ligand interactions. Based on our assessment, students enjoy the exercises, become more interested in bioinformatics and computer modeling, and demonstrate an increase in content knowledge relevant to the topics

Clinical variants of the native class d beta-lactamase of acinetobacter baumannii pose an emerging threat through ıncreased hydrolytic activity against carbapenems

The threat posed by the chromosomally encoded class D beta-lactamase of Acinetobacter baumannii (OXA-51/66) has been unclear, in part because of its relatively low affinity and turnover rate for carbapenems. Several hundred clinical variants of OXA51/66 have been reported, many with substitutions of active-site residues. We determined the kinetic properties of OXA-66 and five clinical variants with respect to a wide variety of beta-lactam substrates. The five variants displayed enhanced activity against carbapenems and in some cases against penicillins, late-generation cephalosporins, and the monobactam aztreonam. Molecular dynamics simulations show that in OXA-66, P130 inhibits the side-chain rotation of I129 and thereby prevents doripenem binding because of steric clash. A single amino acid substitution at this position (P130Q) in the variant OXA-109 greatly enhances the mobility of both I129 and a key active-site tryptophan (W222), thereby facilitating carbapenem binding. This expansion of substrate specificity represents a very worrisome development for the efficacy of beta-lactams against this troublesome pathogen.HHS \ NIH \ National Institute of Allergy and Infectious Diseases (NIAID):1R15AI082416 National Science Foundation (NSF): CNS-122829

Simulations of Antibiotic Resistance: Mining the Trajectory Data

Simulations of Antibiotic Resistance: Mining the Trajectory Data Computer simulations of enzyme dynamics reveal details of the mechanisms through which beta-lactamases select and inactivate the antibiotic molecules. We have a great need to design antibiotics that will withstand the emerging bacterial resistance and long time-scale data will help us with that challenge

Factors Determining Carbapenemase Activity in the OXA Family of ²-Lactamases

Bacterial resistance to antibiotic therapies, especially to ²-lactams is a growing health-care problem. Resistance mediated by class D ²-lactamases has been both the least studied, and most rapidly expanding in the past decade. Of particular clinical concern is the emergence of class D enzymes with the ability to hydrolyze the newest family of ²-lactams: the carbapenems. Class D ²-lactamases are extremely diverse in terms of sequence and hydrolytic profiles, and it remains unclear what factors determine multispecificity in general, and carbapenemase activity in particular. Recent studies have revealed the importance of the ²5-²6 loop in acquiring carbapenemase activity1. Here we present a combined experimental and computational study of the effects that several point mutations in the OXA-24s ²5-²6 loop have on the enzyme catalytic profile. Site-directed mutagenesis and kinetic assays indeed show significant changes in the catalytic profiles of the mutant enzymes. We have employed several computational techniques, namely sequence and motif analysis, Molecular Dynamics simulations and covariance analysis based on the Anisotropic Network Model in order to determine the impact of M223A, G224D and P227S mutations on the dynamics of the OXA-24 active site. We show here that the mutations affect the dynamics of the catalytic site, specifically the carboxylated lysine residue and its hydrogen bonding network within the binding pocket. Multiple sequence alignment and motif analysis show distinct patterns of the ²5-²6 loop sequence variation in different subgroups of OXA carbapenemases. These data will help correlate the sequence traits of OXA carbapenemases to their mechanism of substrate selection and hydrolysis

Investigating the Effects of BIBR1532 and Related Analogs on Telomerase Activity in Human Prostate Cancer Cells

Unlimited cellular proliferation of cancer cells is coupled with the maintenance of telomeres in DNA. Telomerase, the enzyme that re-extends telomeres, has become an attractive target for new cancer therapeutics. BIBR1532, a mixed-type non-competitive inhibitor of telomerase, has been shown to cause growth arrest in tumor cells. Here, we tested BIBR1532 and five synthetic analogues (WS6-48, WS4-43A, WS5-29, WS7-6, WS8-3) for anti-proliferative activity on metastatic prostate cancer cells. Preliminary results indicate these compounds are highly active against proliferation. Their effects on inhibiting telomerase activity directly were quantified using a telomere repeat amplification protocol assay. Newly developed analogues are under preliminary testing to determine their effect upon telomerase. Available x-ray structures of telomerase domains are also being explored to asses putative binding sites and affinities. Further studies will assess the effect these compounds have on other metastatic cancer cell lines

Autoantibodies, C-reactive protein, erythrocyte sedimentation rate and serum cytokine profiling in monitoring of early treatment

Introduction: Currently used clinical scale and laboratory markers to monitor patients with early rheumatoid arthritis (RA) seem to be not sufficient. It has been demonstrated that disease- related cytokines may be elevated very early in RA development and cytokines are considered as the biomarkers potentially useful for RA monitoring. Material and methods : The group of patients with undifferentiated arthritis (UA) developing RA (UA→RA) was identified from a total of 121 people with arthralgia. UA→RA (n = 16) and healthy control (n = 16) subjects underwent clinical and laboratory evaluation, including acute phase reactants (APRs) and autoantibodies. Cytokines IFN-γ, IL-10, TNF, IL-17A, IL-6, IL-1b, IL-2 in sera were assayed using flow cytometric bead array test. Results : 34.5% of patients with UA developed RA. DAS28 reduced as early as 3 months after initiation of treatment. No DAS28 difference between groups of autoantibody (RF, anti-CCP, ANA-HEp-2) -positive and -negative patients was observed, however, comparing groups of anti-CCP and RF-double negative and -double positive patients, the trend of sooner clinical improvement was visible in the second abovementioned group. After the treatment introduction, the ESR level reduced significantly, while CRP level reduction was not significant. Serum cytokine levels of IL-10, IL-6 and IL-17A reduced after 6 months since introduction of treatment. The positive correlations between ESR, CRP and specific cytokine levels were observed. Conclusions : The autoantibody and APR profile is poorly connected with the RA course. The serum cytokine profile change in the course of RA and may be potentially used for optimization of RA monitoring

A Distal Disulfide Bridge in OXA‑1 β‑Lactamase Stabilizes the Catalytic Center and Alters the Dynamics of the Specificity Determining Ω Loop

Widespread antibiotic resistance, particularly when mediated by broad-spectrum β-lactamases, has major implications for public health. Substitutions in the active site often allow broad-spectrum enzymes to accommodate diverse types of β-lactams. Substitutions observed outside the active site are thought to compensate for the loss of thermal stability. The OXA-1 clade of class D β-lactamases contains a pair of conserved cysteines located outside the active site that forms a disulfide bond in the periplasm. Here, the effect of the distal disulfide bond on the structure and dynamics of OXA-1 was investigated via 4 μs molecular dynamics simulations. The results reveal that the disulfide promotes the preorganized orientation of the catalytic residues and affects the conformation of the functionally important Ω loop. Furthermore, principal component analysis reveals differences in the global dynamics between the oxidized and reduced forms, especially in the motions involving the Ω loop. A dynamical network analysis indicates that, in the oxidized form, in addition to its role in ligand binding, the KTG family motif is a central hub of the global dynamics. As activity of OXA-1 has been measured only in the reduced form, we suggest that accurate assessment of its functional profile would require oxidative conditions mimicking periplasm

Intra-/Intermolecular crosslinks identified by mass spectrometric studies of higher order oligomeric GlyBP bands.

<p>*the ΔMass is the maximum observed ppm difference between theoretical and observed m/z over N, the number of times this m/z peak was observed in 10 independent experiments. See <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0102571#pone-0102571-g005" target="_blank"><b>Fig. 5</b></a> for corresponding assigned peak in representative MALDI-TOF spectrum.</p><p>Unique bands found in higher order oligomeric GlyBP bands (absent in monomeric bands) and assigned as intermolecular crosslinks are <i>italicized</i>, with <b><i>bold</i></b> indicating assignments that <b><i>cannot</i></b> be assigned as an intramolecular crosslinks.</p