Residual Dipolar-Coupling-Based Conformational Comparison of Noncovalent Ubiquitin Homodimer with Covalently Linked Diubiquitin.

Although the conformation of the polymer chain of Ubiquitin (Ub) mainly depends on the type of isopeptide linkage connecting two Ub molecules, the non-covalent (noncovalent) interaction between two Ub molecules within the chain could also tune their conformational preference. Here, we studied the conformation of noncovalently formed Ub dimers in solution using residual dipolar couplings (RDCs). Comparing the RDC derived alignment tensor of the noncovalently formed dimer with the two most abundant (K11 and K48) covalent linked Ub dimers revealed that the conformation of K11 linked and noncovalent Ub dimers were similar. Between the various NMR and crystal structures of K11 linked Ub dimers, RDC tensor analysis showed that the structure of K11 linked dimer crystalized at neutral pH is similar to noncovalent dimer. Analogous to the experimental study, the comparison of predicted order matrix of various covalent Ub dimers with that of the experimentally determined order matrix of noncovalent Ub dimer also suggests that the conformation of K11 linked dimers crystalized at neutral pH is similar to the noncovalent dimer.

GTP-binding facilitates EB1 recruitment onto microtubules by relieving its auto-inhibition.

Microtubule plus end-binding protein, EB1 is a key regulator of microtubule dynamics. Auto-inhibitory interaction in EB1 has previously been shown to inhibit its ability to bind to microtubules and regulate microtubule dynamics. However, the factors that promote its microtubule regulatory activity by over-coming the auto-inhibition are less known. Here, we show that GTP plays a critical role in promoting the microtubule-targeting activity of EB1 by suppressing its auto-inhibition. Our biophysical data demonstrate that GTP binds to EB1 at a distinct site in its conserved N-terminal domain. Detailed analyses reveal that GTP-binding suppresses the intra-molecular inhibitory interaction between the globular N-terminus and the C-terminal coiled-coil domain. We further show that mutation of the GTP-binding site residues in N-terminus weakens the affinity for GTP, but also for the C-terminus, indicating overlapping binding sites. Confocal imaging and biochemical analysis reveal that EB1 localization on the microtubules is significantly increased upon mutations of the GTP-binding site residues. The results demonstrate a unique role of GTP in facilitating EB1 interaction with the microtubules by relieving its intra-molecular inhibition. They also implicate that GTP-binding may regulate the functions of EB1 on the cellular microtubules.

Direct Observation of Aggregation-Induced Backbone Conformational Changes in Tau Peptides.

In tau proteins, the hexapeptides in the R2 and R3 repeats are known to initiate tau fibril formation, which causes a class of neurodegenerative diseases called the taupathies. We show that in R3, in addition to the presence of the hexapeptides, the correct turn conformation upstream to it is also essential for producing prion-like fibrils that are capable of propagation. A time-dependent NMR aggregation assay of a slow fibril forming R3-S316P peptide revealed a trans to cis equilibrium shift in the peptide-bond conformation preceding P316 during the growth phase of the aggregation process. S316 was identified as the key residue in the turn that confers templating capacity on R3 fibrils to accelerate the aggregation of the R3-S316P peptide. These results on the specific interactions and conformational changes responsible for tau aggregation could prove useful for developing an efficient therapeutic intervention in Alzheimer's disease.

Pregabalin Abuse amongst Opioid Substitution Treatment Patients.

Pregabalin (Lyrica®) is used in treating epilepsy, nerve pain and anxiety. Pregabalin was initially thought to have a low misuse potential however there are emerging reports of Pregabalin being abused. A study was commenced at the National Drug Treatment Centre's (NDTC) Drug Analysis Laboratory to determine the level of usage of Pregabalin within the addiction services population in Ireland. A total of 498 urine samples representing samples from 440 individual opioid substitution patients, initially screened by immunoassay for drugs of abuse, were subjected to further analysis for Pregabalin by Liquid Chromatography/Mass Spectrometry (LC/MS). Of 440 patients tested, 39 tested positive for Pregabalin (9.2%). Only 10 patients from this group were prescribed this drug to our knowledge thus giving an estimated rate of misuse of 7.0%. Other drugs detected in the Pregabalin positive patients were Opiates (31.8%), Cocaine (11.4%), Benzodiazepines (79.5%) and Cannabis (77.8%). Our study confirms that Pregabalin abuse is taking place amongst the addiction services population. We believe that misuse of this prescription drug is a serious emerging issue which should be monitored carefully.

Molecular Balances Based on Aliphatic CH-π and Lone-Pair-π Interactions.

CH···π and lone-pair···π interactions are estimated for a series of conformationally dynamic bicyclic N-aryliimides. On the basis of their strengths and mutual synergy/competition, the molecules prefer a folded/unfolded conformation. Calculations suggest strategies to selectively isolate the folded form by increasing the strength of the attractive CH···π interaction or removing the lone-pair···π repulsion. While the barrier for the folded ⇄ unfolded transformation is too large to conformationally lock the molecules in either of the conformers, the dynamics for hopping of the alkyl group across rings and tumbling over the rings are found to be facile in the folded conformation.

Role of the pharmacist in pre-exposure chemoprophylaxis (PrEP) therapyfor HIV prevention / Papel del farmacéutico en la quimioprofilaxis pre-exposición para prevención del VIH

With a global estimate of 2.5 million new infections of HIV occurring yearly, discovering novel methods to help stem the spread of the virus is critical. The use of antiretroviral chemoprophylaxis for preventing HIV after accidental or occupational exposure and in maternal to fetal transmission has become a widely accepted method to combat HIV. Based on this success, pre-exposure chemoprophylaxis (PrEP) is being explored in at-risk patient populations such as injecting drug users, female sex workers and men who have sex with men. This off-label and unmonitored use has created a need for education and intervention by pharmacists and other healthcare professionals. Pharmacists should educate themselves on PrEP and be prepared to counsel patients about their means of obtaining it (e.g. borrowing or sharing medications and ordering from disreputable Internet pharmacies). They should also be proactive about medication therapy management in these patients due to clinically important drug interactions with PrEP medications. Only one trial exploring the safety and efficacy of tenofovir as PrEP has been completed thus far. However, five ongoing trials are in various stages and two additional studies are scheduled for the near future. Unfortunately, studies in this arena have met with many challenges that have threatened to derail progress. Ethical controversy surrounding post-trial care of participants who seroconvert during studies, as well as concerns over emerging viral resistance and logistical site problems, have already halted several PrEP trials. Information about these early trials has already filtered down to affected individuals who are experimenting with this unproven therapy as an 'evening before pill'. The potential for PrEP is promising; however, more extensive trials are necessary to establish its safety and efficacy. Pharmacists are well-positioned to play a key role in helping patients make choices about PrEP, managing their therapy, and developing policy with an eye towards the future (AU)

Con una estimación global de 2,5 millones de nuevas infecciones de VIH cada año, es crítico descubrir nuevos métodos para ayudar a detener la extensión del virus. El uso de quimioprofilaxis antirretroviral para prevención del VIH después de una exposición accidental u ocupacional y en transmisión materno-fetal se ha convertido en un método comúnmente aceptado para combatir el VIH. Basando en este éxito, se está explorando la quimioprofilaxis pre-exposición (PrEP) en poblaciones de riesgo tales como drogadictos intravenosos, trabajadoras sexuales, y hombres que tiene sexo con hombres. Este uso no autorizado y no vigilado ha creado la necesidad de educación e intervención de farmacéuticos y otros profesionales de la salud. Los farmacéuticos deberían formarse sobre la PrEP y estar preparados para aconsejar a los pacientes sobre los medios de obtenerla (p.e. prestándose o compartiendo medicaciones y comprándola en farmacias de confianza en Internet). También deberían ser proactivos en el seguimiento de estos pacientes debido a las importantes interacciones de los medicamentos PrEP. Solo se ha completado un ensayo que exploró la seguridad y la eficacia del tenofovir como PrEP. Sin embargo, están programados 5 ensayos en marcha en diversos estadios, y dos estudios adicionales en un futuro próximo. Desafortunadamente, los estudios en este campo se han encontrado con muchos obstáculos que han amenazado interrumpir su progreso. La controversia ética sobre la atención post-ensayo de los pacientes que se seroconvierten durante el estudio, así como las preocupaciones sobre la emergente resistencia viral y los problemas logísticos, ya han interrumpido varios ensayos de PrEP. La información de estos primeros ensayos se ha filtrado e los individuos afectados que están experimentando con este tratamiento no probado como una 'píldora de la tarde anterior'. El potencial de la PrEP es prometedor, sin embargo se necesitan ensayos más extensos para establecer su seguridad y eficacia. Los farmacéuticos están bien posicionaos para jugar un papel importante ayudando a los pacientes a tomar mejores decisiones sobre la PrEP, gestionando su tratamiento, y desarrollando políticas con un ojo en el futuro (AU)

Role of the pharmacist in pre-exposure chemoprophylaxis (PrEP) therapy for HIV prevention.

With a global estimate of 2.5 million new infections of HIV occurring yearly, discovering novel methods to help stem the spread of the virus is critical. The use of antiretroviral chemoprophylaxis for preventing HIV after accidental or occupational exposure and in maternal to fetal transmission has become a widely accepted method to combat HIV. Based on this success, pre-exposure chemoprophylaxis (PrEP) is being explored in at-risk patient populations such as injecting drug users, female sex workers and men who have sex with men. This off-label and unmonitored use has created a need for education and intervention by pharmacists and other healthcare professionals. Pharmacists should educate themselves on PrEP and be prepared to counsel patients about their means of obtaining it (e.g. borrowing or sharing medications and ordering from disreputable Internet pharmacies). They should also be proactive about medication therapy management in these patients due to clinically important drug interactions with PrEP medications. Only one trial exploring the safety and efficacy of tenofovir as PrEP has been completed thus far. However, five ongoing trials are in various stages and two additional studies are scheduled for the near future. Unfortunately, studies in this arena have met with many challenges that have threatened to derail progress. Ethical controversy surrounding post-trial care of participants who seroconvert during studies, as well as concerns over emerging viral resistance and logistical site problems, have already halted several PrEP trials. Information about these early trials has already filtered down to affected individuals who are experimenting with this unproven therapy as an "evening before pill". The potential for PrEP is promising; however, more extensive trials are necessary to establish its safety and efficacy. Pharmacists are well-positioned to play a key role in helping patients make choices about PrEP, managing their therapy, and developing policy with an eye towards the future.

Acceptability and use of sexual barrier products and lubricants among HIV-seropositive Zambian men.

This study assessed the acceptability and preference for sexual barrier and lubricant products among men in Zambia following trial and long-term use. It also examined the role of men's preferences as facilitators or impediments to product use for HIV transmission reduction within the Zambian context. HIV-seropositive and -serodiscordant couples were recruited from HIV voluntary counseling and testing centers in Lusaka between 2003 and 2006; 66% of those approached agreed to participate. HIV seropositive male participants participated in a product exposure group intervention (n = 155). Participants were provided with male and female condoms and vaginal lubricants (Astroglide [BioFilm, Inc., Vista, CA] & KY gels [Johnson & Johnson, Langhorne, PA], Lubrin suppositories [Kendwood Therapuetics, Fairfield, NJ]) over three sessions; assessments were conducted at baseline, monthly over 6 months and at 12 months. At baseline, the majority of men reported no previous exposure to lubricant products or female condoms and high (79%) levels of consistent male condom use in the last 7 days. Female condom use increased during the intervention, and male condom use increased at 6 months and was maintained over 12 months. The basis for decisions regarding lubricant use following product exposure was most influenced by a preference for communicating with partners; participant preference for lubricant products was distributed between all three products. Results illustrate the importance of development of a variety of products for prevention of HIV transmission and of inclusion of male partners in interventions to increase sexual barrier product use to facilitate barrier acceptability and use in Zambia.

Both under-nutrition and obesity increase morbidity following liver transplantation.

Malnutrition is common in patients awaiting liver transplantation and may contribute to operative and post-operative mortality, although this is controversial. We assessed the pre-operative nutritional status of 87 patients and the impact this had on mortality and morbidity following liver transplantation for chronic liver disease. Thirty six per cent of patients had more than 10% loss of body weight prior to transplantation. Nutritional depletion, considered present if triceps skin fold thickness or mid-arm muscle circumference were < 5th percentile, was present in 17% and 15% of the total group respectively. Patients whose pre-operative body weights were < 90% of their ideal body weight (IBW) had a longer hospital stay (p = 0.001) and required longer post-operative ventilatory support (p = 0.033). This group also required significantly more treatment with intravenous antibiotics (p = 0.001) suggesting an increased incidence of infective complications. Patients who were obese pre-operatively (body mass index > 30Kg/m2) also required a longer period in high dependency (p = 0.0003). No individual nutritional variable correlated with mortality. In the Irish population undergoing liver transplantation, we found a relatively low prevalence of malnutrition in comparison with other studies. Both under- nutrition and obesity significantly affected morbidity and length of hospital stay post-transplant, although no individual nutritional variable predicted survival post transplant.