The C-terminus of ICln is natively disordered but displays local structural preformation

ICln is a vital, ubiquitously expressed protein with roles in cell volume regulation, angiogenesis, cell morphology, activation of platelets and RNA processing. In previous work we have determined the 3D structure of the N-terminus of ICln (residues 1-159), which folds into a PH-like domain followed by an unstructured region (residues H134 - Q159) containing protein-protein interaction sites. Here we present sequence-specific resonance assignments of the C-terminus (residues Q159 - H235) of ICln by NMR, and show that this region of the protein is intrinsically unstructured. By applying (13)Cα- (13)Cβ secondary chemical shifts to detect possible preferences for secondary structure elements we show that the C-terminus of ICln adopts a preferred α-helical organization between residues E170 and E187, and exists preferentially in extended conformations (β-strands) between residues D161 to Y168 and E217 to T223.     

When less is more: A simple predictive model for repeated prostate biopsy outcomes

ObjectivesTo present a new predictive model for repeated prostate biopsy outcomes. Several practical problems are described that arise when searching for a proper model among those that already exist. A new model is developed with only two explanatory variables and a simple graphical output.MethodsThis is a retrospective cohort study based on data collected from December 2006 to June 2011 at the Clinic of Urology of the University Hospital in Olomouc, Czech Republic. The cohort consists of 221 patients who underwent the first repeated biopsy after an initial biopsy with a negative outcome. All patients had prostate-specific antigen (PSA) levels between 1.5 and 16.5 ng/mL and a prostate volume not greater than 100 mL. A logistic regression model was fitted.ResultsOf the 221 patients, 29 (13%) were diagnosed with prostate cancer on the repeated biopsy. The final model includes the PSA level and the transitory zone volume as predictors. Its accuracy is 76.4%. The cut-off point of 0.0687 in the predicted positive repeated biopsy outcome assures 95% sensitivity and prevents 42% of unnecessary biopsies.ConclusionsThe accuracy of the model is comparable to that of more complex models (with more than two predictors) published in the literature. The model includes only two routinely measured variables, and hence it is accessible for a wide range of practitioners. The simple graphical outcome makes the model even more attractive.     

Prospective Multicenter Study of Community-Associated Skin and Skin Structure Infections due to Methicillin-Resistant Staphylococcus aureus in Buenos Aires,Argentina

Background

Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) is now the most common cause of skin and skin structure infections (SSSI) in several world regions. In Argentina prospective, multicenter clinical studies have only been conducted in pediatric populations.

Objective

Primary: describe the prevalence, clinical and demographic characteristics of adult patients with community acquired SSSI due to MRSA; secondary: molecular evaluation of CA-MRSA strains. Patients with MRSA were compared to those without MRSA.

Materials and Methods

Prospective, observational, multicenter, epidemiologic study, with molecular analysis, conducted at 19 sites in Argentina (18 in Buenos Aires) between March 2010 and October 2011. Patients were included if they were ≥14 years, were diagnosed with SSSI, a culture was obtained, and there had no significant healthcare contact identified. A logistic regression model was used to identify factors associated with CA-MRSA. Pulse field types, SCCmec, and PVL status were also determined.

Results

A total of 311 patients were included. CA-MRSA was isolated in 70% (218/311) of patients. Clinical variables independently associated with CA-MRSA were: presence of purulent lesion (OR 3.29; 95%CI 1.67, 6.49) and age <50 years (OR 2.39; 95%CI 1.22, 4.70). The vast majority of CA-MRSA strains causing SSSI carried PVL genes (95%) and were SCCmec type IV. The sequence type CA-MRSA ST30 spa t019 was the predominant clone.

Conclusions

CA-MRSA is now the most common cause of SSSI in our adult patients without healthcare contact. ST30, SCCmec IV, PVL+, spa t019 is the predominant clone in Buenos Aires, Argentina.     

ImmunoPET and biodistribution with human epidermal growth factor receptor 3 targeting antibody 89Zr-RG7116

The humanized monoclonal antibody with high affinity for the human epidermal growth factor receptor (HER) 3, RG7116, is a glycoengineered, IgG1 class antibody. By labeling RG7116 with zirconium-89 (⁸⁹Zr) we aimed to visualize in vivo HER3 expression and study the biodistribution of this antibody in human tumor-bearing mice. Biodistribution of ⁸⁹Zr-RG7116 was studied in subcutaneously xenografted FaDu tumor cells (HER3-positive). Dose-dependency of ⁸⁹Zr-RG7116 organ distribution and specific tumor uptake was assessed by administering doses ranging from 0.05 to 10 mg/kg RG7116 to SCID/Beige mice. Biodistribution was analyzed at 24 and 144 h after injection. MicroPET imaging was performed at 1, 3, and 6 days after injection of 1.0 mg/kg ⁸⁹Zr-RG7116 in the FaDu, H441, QG-56 and Calu-1 xenografts with varying HER3 expression. The excised tumors were analyzed for HER3 expression. Biodistribution analyses showed a dose- and time-dependent ⁸⁹Zr-RG7116 tumor uptake in FaDu tumors. The highest tumor uptake of ⁸⁹Zr-RG7116 was observed in the 0.05 mg/kg dose group with 27.5%ID/g at 144 h after tracer injection. MicroPET imaging revealed specific tumor uptake of ⁸⁹Zr-RG7116 in FaDu and H441 models with an increase in tumor uptake over time. Biodistribution data was consistent with the microPET findings in FaDu, H441, QG56 and Calu-1 xenografts, which correlated with HER3 expression levels. In conclusion, ⁸⁹Zr-RG7116 specifically accumulates in HER3 expressing tumors. PET imaging with this tracer provides real-time non-invasive information about RG7116 distribution, tumor targeting and tumor HER3 expression levels.     

Control of Oxidative Damage in Rheumatoid Arthritis By Gold(I)-Thiolate Drugs

The roles of anti-arthritic gold(I)-thiolate drugs such as disodium aurothiomalate ('Myocrisin') in the modulation or promotion of oxygen radical-mediated oxidative damage in vivo ate reviewed. In particular, the precise molecular mechanisms by which these novel second-line agents exert their therapeutic effects are discussed in terms of (i) the direct and indirect control of enzymes involved in the generation or scavenging of reactive oxygen speices (ROS) such as superoxide ion, hydrogen peroxide and hydroxyl radical, (ii) the protection of proteins and relevant enzyme systems against attack by ROS and (iii) their direct involvement in the production (at appropriate 'target' sites) or scavenging of ROS in vivo. In addition, the role of the orally-effective gold(I)-phosphine complex auranofin in the control of oxidative damage in rheumatoid arthritis is also discussed.     

Synthesis of enantiomeric pureCis-dichloroplatinum(II) amino acid complexes

The reaction of potassium tetrachloroplatinate(II) with six representative sulfurcontaining amino acids, namely,d- andl-cysteine,d- andl-methionine and its methyl ester hydrochloride gives the corresponding enantiomerically purecis-dichloroplatinum(II) complexes. This represents the first reported series of well-characterized enantiomerically pure platinum(II) complexes for bothd- andl-amino acids. The spectroscopic properties, including IR,¹H-NMR, and¹³C NMR, of these complexes and their configuration are discussed.     

The temporal order of replication of murine immunoglobulin heavy chain constant region sequences corresponds to their linear order in the genome.

The time of replication during the S phase in a murine erythroleukemia (MEL) cell line was determined for immunoglobulin heavy chain constant region C alpha, C gamma 2b and C mu sequences whose boundaries are defined by EcoR1 restriction endonuclease sites (EcoR1 segments). Logarithmically growing cultures of MEL cells with an S phase of about 7.5 hours were pulse labelled with 20 micrograms/ml of 5-bromodeoxyuridine (BUdR). The cells were then fractionated by centrifugal elutriation into 10-12 distinct populations containing cells in different stages of the cell cycle. Flow microfluorimetric (FMF) analysis of DNA content, measurements of cell volume and autoradiography after 3H-thymidine pulse labelling were used to determine position in the cell cycle. Fractions were pooled to represent four selected intervals of S in which BU-DNA was synthesized for 2.5 hrs or less. Newly replicated DNA which had incorporated BUdR into one strand was isolated, cleaved with EcoR1, and separated on neutral Cs2S04 gradients. Equal amounts of BU-DNA replicated during these four intervals of S were electrophoresed in 0.8% agarose gels, transferred to diazotized aminobenzyloxymethyl paper and hybridized with 32p probes containing the C alpha, C gamma 2b and C mu genes and flanking sequences. The relative amounts of segments replicated were assessed by quantitation of the appropriate bands on the autoradiograms by microdensitometry. The results indicate that the 2.8 kb C alpha, 6.6 kb C gamma 2b and 12 kb C mu EcoR1 segments in these MEL cells replicated during defined intervals of the first half of the S phase. The order of replication of these EcoR1 segments as the cells proceeded through S was C alpha, C gamma 2b, C mu, corresponding to the linear order of the genes determined by restriction endonuclease mapping.     

Induction of peroxisome proliferation and increase of catalase activity in yeast,Candida albicans,by cadmium

The effects of cadmium on the growth rate, catalase activity, and peroxisome proliferation in yeast,Candida albicans, were evaluated. The yeast growth was markedly inhibited by 1 mM cadmium at the initial hours. The toxic effect of cadmium on the cell growth persisted. The catalase activity of the cells treated with 1 mM Cd²⁺ first decreased, and then rose at 24 h to about 2.6 times that of the controls. The average number of peroxisomes per cell in the yeast treated with 1 mM Cd²⁺ was about sixfold higher than the control groups. The proliferation of peroxisomes and the increase of catalase activity following cadmium toxicity gives credence to the hypothesis that cadmium toxicity is related to its potential to induce oxidative stress in cells.