Controlled assembly of SNAP-PNA-fluorophore systems on DNA templates to produce fluorescence resonance energy transfer

The SNAP protein is a widely used self-labeling tag that can be used for tracking protein localization and trafficking in living systems. A model system providing controlled alignment of SNAP-tag units can provide a new way to study clustering of fusion proteins. In this work, fluorescent SNAP-PNA conjugates were controllably assembled on DNA frameworks forming dimers, trimers, and tetramers. Modification of peptide nucleic acid (PNA) with the O6-benzyl guanine (BG) group allowed the generation of site-selective covalent links between PNA and the SNAP protein. The modified BG-PNAs were labeled with fluorescent Atto dyes and subsequently chemo-selectively conjugated to SNAP protein. Efficient assembly into dimer and oligomer forms was verified via size exclusion chromatography (SEC), electrophoresis (SDS-PAGE), and fluorescence spectroscopy. DNA directed assembly of homo- and hetero-dimers of SNAP-PNA constructs induced homo- and hetero-FRET, respectively. Longer DNA scaffolds controllably aligned similar fluorescent SNAP-PNA constructs into higher oligomers exhibiting homo-FRET. The combined SEC and homo-FRET studies indicated the 1:1 and saturated assemblies of SNAP-PNA-fluorophore:DNA formed preferentially in this system. This suggested a kinetic/stoichiometric model of assembly rather than binomially distributed products. These BG-PNA-fluorophore building blocks allow facile introduction of fluorophores and/or assembly directing moieties onto any protein containing SNAP. Template directed assembly of PNA modified SNAP proteins may be used to investigate clustering behavior both with and without fluorescent labels which may find use in the study of assembly processes in cells

A Prospective Pilot Study to Identify a Myocarditis Cohort who may Safely Resume Sports Activities 3 Months after Diagnosis

International cardiovascular society recommendations to return to sports activities following acute myocarditis are based on expert consensus in the absence of prospective studies. We prospectively enrolled 30 patients with newly diagnosed myocarditis based on clinical parameters, laboratory measurements and cardiac magnetic resonance imaging with mildly reduced or preserved left ventricular ejection fraction (LVEF) with a follow-up of 12 months. Cessation of physical activity was recommended for 3 months. The average age was 35 (19-80) years with 73% male patients. One case of non-sustained ventricular tachycardia was recorded during 48-h-Holter electrocardiogram. Except for this case, all patients were allowed to resume physical exercise after 3 months. At 6- (n = 26) and 12-month (n = 19) follow-up neither cardiac events nor worsening LVEF were recorded. The risk of cardiac events at 1 year after diagnosis of myocarditis appears to be low after resumption of exercise after 3 months among patients who recover from acute myocarditis

Has there been any structural convergence in the transmission of European monetary policies?

Time-frequency analysis, Transmission mechanism, Convergence, Monetary policy, C22, C29,

Virome Sequencing in Patients With Myocarditis

BACKGROUND Polymerase chain reaction analyses of cardiac tissues have detected viral sequences in up to 67% of cases of myocarditis. However, viruses have not been implicated in giant cell myocarditis (GCM). Furthermore, efforts to detect viruses implicated in myocarditis have been unsuccessful in more accessible samples such as peripheral blood. METHODS We used Virome Capture Sequencing for Vertbrate Viruses (VirCapSeq-VERT), a method that simultaneously screens for all known vertebrate viruses, to investigate viruses in 33 patients with myocarditis. We investigated peripheral blood mononuclear cells (n=24), plasma (n=27), endomyocardial biopsies (n=2), and cardiac tissue samples from explanted hearts (n=13). RESULTS Nine patients (27%) had GCM and 4 patients (13%) had fulminant myocarditis. We found the following viruses in the blood of patients with myocarditis: Epstein Barr virus (n=11, 41%), human pegivirus (n=1, 4%), human endogenous retrovirus K (n=27, 100%), and anellovirus (n=15, 56%). All tissue samples from fulminant myocarditis (n=2) and GCM (n=13) contained human endogenous retrovirus K. CONCLUSIONS No nucleic acids from viruses previously implicated in myocarditis or other human illnesses were detected in relevant amounts in cardiac tissue samples from GCM or in blood samples from other types of myocarditis. These findings do not exclude a role for viral infection in GCM but do suggest that if viruses are implicated, the mechanism is likely to be indirect rather than due to cytotoxic infection of myocardium