A Smartphone-based Decision Support Tool Improves Test Performance Concerning Application of the Guidelines for Managing Regional Anesthesia in the Patient Receiving Antithrombotic or Thrombolytic Therapy

BACKGROUND: The American Society of Regional Anesthesia and Pain Medicine (ASRA) consensus statement on regional anesthesia in the patient receiving antithrombotic or thrombolytic therapy is the standard for evaluation and management of these patients. The authors hypothesized that an electronic decision support tool (eDST) would improve test performance compared with native physician behavior concerning the application of this guideline. METHODS: Anesthesiology trainees and faculty at 8 institutions participated in a prospective, randomized trial in which they completed a 20-question test involving clinical scenarios related to the ASRA guidelines. The eDST group completed the test using an iOS app programmed to contain decision logic and content of the ASRA guidelines. The control group completed the test by using any resource in addition to the app. A generalized linear mixed-effects model was used to examine the effect of the intervention. RESULTS: After obtaining institutional review board's approval and informed consent, 259 participants were enrolled and randomized (eDST = 122; control = 137). The mean score was 92.4 ± 6.6% in the eDST group and 68.0 ± 15.8% in the control group (P < 0.001). eDST use increased the odds of selecting correct answers (7.8; 95% CI, 5.7 to 10.7). Most control group participants (63%) used some cognitive aid during the test, and they scored higher than those who tested from memory alone (76 ± 15% vs. 57 ± 18%, P < 0.001). There was no difference in time to completion of the test (P = 0.15) and no effect of training level (P = 0.56). CONCLUSIONS: eDST use improved application of the ASRA guidelines compared with the native clinician behavior in a testing environment

Crystal structures of p120RasGAP N-terminal SH2 domain in its apo form and in complex with a p190RhoGAP phosphotyrosine peptide.

The Rho and Ras pathways play vital roles in cell growth, division and motility. Cross-talk between the pathways amplifies their roles in cell proliferation and motility and its dysregulation is involved in disease pathogenesis. One important interaction for cross-talk occurs between p120RasGAP (RASA1), a GTPase activating protein (GAP) for Ras, and p190RhoGAP (p190RhoGAP-A, ARHGAP35), a GAP for Rho. The binding of these proteins is primarily mediated by two SH2 domains within p120RasGAP engaging phosphorylated tyrosines of p190RhoGAP, of which the best studied is pTyr-1105. To better understand the interaction between p120RasGAP and p190RhoGAP, we determined the 1.75 Å X-ray crystal structure of the N-terminal SH2 domain of p120RasGAP in the unliganded form, and its 1.6 Å co-crystal structure in complex with a synthesized phosphotyrosine peptide, EEENI(p-Tyr)SVPHDST, corresponding to residues 1100-1112 of p190RhoGAP. We find that the N-terminal SH2 domain of p120RhoGAP has the characteristic SH2 fold encompassing a central beta-sheet flanked by two alpha-helices, and that peptide binding stabilizes specific conformations of the βE-βF loop and arginine residues R212 and R231. Site-directed mutagenesis and native gel shifts confirm phosphotyrosine binding through the conserved FLVR motif arginine residue R207, and isothermal titration calorimetry finds a dissociation constant of 0.3 ± 0.1 μM between the phosphopeptide and SH2 domain. These results demonstrate that the major interaction between two important GAP proteins, p120RasGAP and p190RhoGAP, is mediated by a canonical SH2-pTyr interaction

Correction: Crystal structures of p120RasGAP N-terminal SH2 domain in its apo form and in complex with a p190RhoGAP phosphotyrosine peptide.

[This corrects the article DOI: 10.1371/journal.pone.0226113.]

Purification and SAXS analysis of the integrin linked kinase, PINCH, parvin (IPP) heterotrimeric complex.

The heterotrimeric protein complex containing the integrin linked kinase (ILK), parvin, and PINCH proteins, termed the IPP complex, is an essential component of focal adhesions, where it interacts with many proteins to mediate signaling from integrin adhesion receptors. Here we conduct a biochemical and structural analysis of the minimal IPP complex, comprising full-length human ILK, the LIM1 domain of PINCH1, and the CH2 domain of α-parvin. We provide a detailed purification protocol for IPP and show that the purified IPP complex is stable and monodisperse in solution. Using small-angle X-ray scattering (SAXS), we also conduct the first structural characterization of IPP, which reveals an elongated shape with dimensions 120×60×40 Å. Flexibility analysis using the ensemble optimization method (EOM) is consistent with an IPP complex structure with limited flexibility, raising the possibility that inter-domain interactions exist. However, our studies suggest that the inter-domain linker in ILK is accessible and we detect no inter-domain contacts by gel filtration analysis. This study provides a structural foundation to understand the conformational restraints that govern the IPP complex

SAXS-derived size parameters for IPP_min.

a<p>Determined by Guinier approximation in Primus <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0055591#pone.0055591-Konarev1" target="_blank">[29]</a>.</p>b<p>Determined in AutoGNOM <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0055591#pone.0055591-Petoukhov1" target="_blank">[30]</a>.</p>c<p>Expected molecular weight = 73,625 Da.</p

Structural modeling of IPP_min based on SAXS data.

<p><b>A</b>) Averaged molecular envelope for IPP_min. The approximate envelope dimensions (in Å) are illustrated. The two views are related by 90° rotation. <b>B</b>) The crystal structures of the individual subunits of the IPP_min complex, ILK-ARD/PINCH-1-LIM1 (PDB code: 3F6Q) and ILK-pseudokinase (pKD)/α-parvin-CH2 (PDB code: 3KMU) used in rigid body modeling. ILK is colored magenta, PINCH-1 is green, and α-parvin is blue. <b>C</b>) CORAL <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0055591#pone.0055591-Petoukhov2" target="_blank">[36]</a> rigid body model of IPP_min (ribbons, colored as in <b>B</b>) with the best statistical fit to the experimental data (plotted in <b>D</b>). Overlaid is the averaged molecular envelope. 14 inter-domain dummy residues between the C-terminus of ILK-ARD and the N-terminus of ILK-pKD, in the optimal conformation chosen by CORAL, are depicted as yellow spheres. The distance between the two subunits is 26 Å. <b>D</b>) Fit of the theoretical scattering profile for the rigid body model (red line) with the experimental SAXS data (logarithmic). Residuals for the fit are shown below.</p

SAXS analysis for IPP_min reveals a globular heterotrimeric complex.

<p><b>A</b>) SAXS intensity profiles (logarithmic) for four concentrations of the IPP_min complex. <b>B</b>) Linearity of Guinier plots with manual selection of Guinier region. The <i>R</i>_g values are presented in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0055591#pone-0055591-t001" target="_blank"><b>Table 1</b></a>. Automatic Guinier analysis performed in AutoRG <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0055591#pone.0055591-Konarev1" target="_blank">[29]</a>, which is consistent with the analysis shown here, is presented in the Supporting Information. <b>C</b>) Normalized pair distribution functions P(<i>R</i>) calculated automatically with AutoGNOM <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0055591#pone.0055591-Petoukhov1" target="_blank">[30]</a>. <b>D</b>) Dimensionless Kratky plots support a globular shape.</p

An unstructured linker in ILK connects the N- and C-terminal subunits of IPP.

<p><b>A</b>) Limited trypsin proteolysis of purified IPP_min complex (lanes 2 through 6) supports that the linker in ILK is unstructured. The N-terminal IPP subunit (ILK-ARD/PINCH-1-LIM1, lane 7) and α-parvin-CH2 alone (lane 8) are included for comparison. Molecular weight markers (in kDa) are shown. <b>B</b>) Gel-filtration chromatography of the full-length IPP_min protein (lane 1 from part D) and trypsin proteolyzed subunit fragments (lane 6 from part D) reveals no apparent interaction between the N- and C-terminal subunits of the IPP complex.</p