1H and 13C resonance assignments of a guanine sensing riboswitch’s terminator hairpin

Here we report the nearly complete base assignments and partial sugar assignments of the 35-residue terminator hairpin of the Bacillus subtilisxpt-pbuX-mRNA guanine sensing riboswitch

Nuclear imaging does not have clear added value in patients with low a priori chance of periprosthetic joint infection. A retrospective single-center experience

Background: A low-grade periprosthetic joint infection (PJI) may present without specific symptoms, and its diagnosis remains a challenge. Three-phase bone scintigraphy (TPBS) and white blood cell (WBC) scintigraphy are incorporated into recently introduced diagnostic criteria for PJI, but their exact value in diagnosing low-grade PJI in patients with nonspecific symptoms remains unclear. Methods: In this retrospective study, we evaluated patients with a prosthetic joint of the hip or knee who underwent TPBS and/or WBC scintigraphy between 2009 and 2016 because of nonspecific symptoms. We reviewed and calculated diagnostic accuracy of the TPBS and/or WBC scintigraphy to diagnose or exclude PJI. PJI was defined based on multiple cultures obtained during revision surgery. In patients who did not undergo revision surgery, PJI was ruled out by clinical follow-up of at least 2 years absent of clinical signs of infection based on MSIS 2011 criteria. Results: A total of 373 patients were evaluated, including 340 TPBSs and 142 WBC scintigraphies. Thirteen patients (3.5 %) were diagnosed with a PJI. TPBS sensitivity, specificity, and positive and negative predictive values (PPV, NPV) were 71 %, 65 %, 8 % and 98 %, respectively. Thirty-five percent of TPBS showed increased uptake. Stratification for time intervals between the index arthroplasty and the onset of symptoms did not alter its diagnostic accuracy. WBC scintigraphy sensitivity, specificity, PPV and NPV were 30 %, 90 %, 25 % and 94 %, respectively. Conclusion: Nuclear imaging does not have clear added value in patients with low a priori chance of periprosthetic joint infection

Thermodynamics and NMR studies on Duck, Heron and Human HBV encapsidation signals

Hepatitis B virus (HBV) replication is initiated by binding of its reverse transcriptase (P) to the apical stem-loop (AL) and primer loop (PL) of epsilon, a highly conserved RNA element at the 5′-end of the RNA pregenome. Mutation studies on duck/heron and human in vitro systems have shown similarities but also differences between their P–epsilon interaction. Here, NMR and UV thermodynamic data on AL (and PL) from these three species are presented. The stabilities of the duck and heron ALs were found to be similar, and much lower than that of human. NMR data show that this low stability stems from an 11-nt internal bulge destabilizing the stem of heron AL. In duck, although structured at low temperature, this region also forms a weak point as its imino resonances broaden to disappearance between 30 and 35°C well below the overall AL melting temperature. Surprisingly, the duck- and heron ALs were both found to be capped by a stable well-structured UGUU tetraloop. All avian ALs are expected to adhere to this because of their conserved sequence. Duck PL is stable and structured and, in view of sequence similarities, the same is expected for heron - and human PL

The presence of extracellular matrix degrading metalloproteinases during fetal development of the intervertebral disc

Matrix metalloproteinases (MMPs) regulate connective tissue architecture and cell migration through extracellular matrix (ECM) degradation and are associated with both physiological and pathological processes. Although they are known to play a role in skeletal development, little is known about the role of MMPs in intervertebral disc (IVD) development. Sixteen fetal human lumbar spine segments, obtained at autopsy, were compared with five normal, non-fetal L4–L5 IVDs. Intensity and/or localization of immunohistochemical staining for MMP-1, -2, -3 and -14 were evaluated by three independent observers. MMP-2 production and activation was quantified by gelatin zymography. MMP-1 and -14 were abundantly present in the nucleus pulposus (NP) and notochordal (NC) cells of the fetal IVDs. In non-fetal IVDs, MMP-1 and -14 staining was significantly less intense (p = 0.001 and p < 0.001, respectively). MMP-3 was found in almost the entire IVD with no significant difference from non-fetal IVDs. MMP-2 staining in the NC and NP cells of the fetal IVD was moderate, but weak in the non-fetal IVD. Gelatin zymography showed a negative correlation of age with MMP-2 activity (p < 0.001). MMP-14 immunostaining correlated positively with MMP-2 activity (p = 0.001). For the first time, the presence of MMP-1, -2, -3 and -14 in the fetal human IVD is shown and the high levels of MMP-1, -2 and -14 suggest a role in the development of the IVD. In particular, the gradual decrease in MMP-2 activation during gestation pinpoints this enzyme as key player in fetal development, possibly through activation by MMP-1 and -14

Negative Pressure Incision Management System in the Prevention of Groin Wound Infection in Vascular Surgery Patients

Objectives. Groin wounds following vascular surgery are highly susceptible to healing disturbances, with reported site infections reaching 30%. Negative pressure incision management systems (NPIMS) are believed to positively influence the prevention of surgical wound-healing disturbances (WHD) and surgical site infections (SSI). NPIMS placed directly after closure of the surgical wound is thought to result in fewer infections; we analysed its effect on postoperative wound infections in patients after vascular surgery via the groin. Methods. From May 2012 to March 2013 we included 90 surgical patients; 40 received a NPIMS. All patients with WHDs were labelled and subanalysed for surgical site infection in case of positive microbiological culture. These infections were graded according to Szilagyi. Number of WHDs and SSIs were compared across cohorts. Results. Patient and perioperative characteristics were equal, except for a significantly higher number of emergency procedures among non-NPIMS patients. We found no significant differences in number of WHDs, SSIs, or Szilagyi grades between the two cohorts. Conclusion. The equal number of SSIs across cohorts showed that NPIMS could not reduce the number of surgical site infections after vascular groin surgery

Transcription and translation regulating riboswitches. structure of the guanine sensing riboswitch in its guanine-free and -bound forms and comparison with the SAM Riboswitch type II

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Ligand-induced folding of the guanine-sensing riboswitch is controlled by a combined predetermined–induced fit mechanism

All known guanine-sensing riboswitches regulate gene expression by specifically binding to guanine (G) or related analogs with high affinity to switch off transcription. The aptamers of this class of riboswitches are characterized by three helices (P1–P3), surrounding a central core of phylogenetically conserved nucleotides and a long-range loop–loop interaction. To gain more insight into the switching mechanism, we present here a comparison between the solution-state structures of the G-free and G-bound forms of the guanine aptamer from the xpt-pbuX operon of Bacillus subtilis, as derived from NMR chemical shifts and magnetic-field-induced residual dipolar couplings. The high-resolution NMR analysis shows the G-free aptamer is highly structured with parallel P2 and P3 helices and the long-range loop–loop interaction already present, implying that the structure is largely preformed to bind the ligand. Structural changes upon guanine binding are found to be localized to the central core. In the free state, the G-quadruple interaction and two base pairs of the P1 stem flanking the central core appear to be largely disordered. The ligand thus binds via a combined predetermined–induced fit mechanism, involving a previously unstructured five-residue loop of the J2–3 junction that folds over the ligand. These limited additional interactions within a preorganized setting possibly explain how the aptamer rapidly responds to ligand binding, which is necessary to switch the structural state of the expression platform within a narrow time frame before the RNA polymerase escapes the 5′-UTR

Difftrain: A Novel Approach to a True Spectroscopic Single-Scan Diffusion Measurement

Diffusion-ordered spectroscopy (DOSY) has gained considerable attention over the past decade as a useful tool for calculating diffusion-related parameters or in the analysis of complex (reaction) mixtures. A major drawback of the established methods are the relatively long recording times needed to acquire the spectra, excluding the monitoring of rapidly progressing reactions and (hence) measurements of less stable components. In order to overcome these shortcomings a new pulse sequence, Difftrain, has been developed. The sequence involves stimulated echo attenuation, multilow flip angle excitation, and multiple sampling of the FID during the longitudinal storage. The calculated diffusion parameters obtained by Difftrain were compared with those obtained by the established sequence BPPSTE (bipolar pulse pair stimulated echo) and were in good agreement. For systems with moderate to good signal-to-noise ratios the Difftrain building block yields significant saving in recording time (single-shot acquisition instead of acquiring n-different gradients strengths), thus opening up new applications in nonequilibrium systems and screening of compositions and/or interactions of (larger) compound arrays