Monomer Choice Influences N-Acryloyl Amino Acid Grafter Conversion via Protease Catalysis.

End-capped peptides modified with reactive functional groups on the N-terminus provide a route to prepare peptide-polymer conjugates for a broad range of applications. Unfortunately, current chemical methods to construct modified peptides rely largely on solid-phase peptide synthesis (SPPS), which lacks green preparative characteristics and is costly, thus limiting its applicability to specialty applications such as regenerative medicine. This work evaluates N-terminally modified N-acryloyl-glutamic acid diethyl ester, N-acryloyl-leucine ethyl ester, and N-acryloyl-alanine ethyl ester as grafters and papain as the protease for the direct addition of amino acid ethyl ester (AA-OEt) monomers via protease-catalyzed peptide synthesis (PCPS) and the corresponding formation of N-acryloyl-functionalized oligopeptides in a one-pot aqueous reaction. It was hypothesized that by building N-acryloyl grafters from AA-OEt monomers that are known to be good substrates for papain in PCPS, the corresponding grafters would yield high grafter conversions, high ratio of grafter-oligopeptide to free NH2-oligopeptide, and high overall yield. However, this work demonstrates based on the grafter/monomers studied herein that the dominant factor in N-acryloyl-AA-OEt grafter conversion is the co-monomer used in co-oligomerizations. Computational modeling using Rosetta qualitatively recapitulates the results and provides insight into the structural and energetic bases underlying substrate selectivity. The findings herein expand our knowledge of factors that determine the efficiency of preparing N-acryloyl-terminated oligopeptides by PCPS that could provide practical routes to peptide macromers for conjugation to polymers and surfaces for a broad range of applications.

Emergency Department Operations II: Patient Flow.

Emergency departments have always been busy, but persistent annual increases in volume and rates of admission have led to high levels of boarding patients, adding additional risk to the situation. This article reviews specific areas of risk as patients progress through their care in the emergency department and methods by which to mitigate this risk. Beginning with an overview of the current state, commonly used throughput metrics are reviewed before proceeding to a discussion of best practice strategies to decrease risk exposure at input, throughput, and output phases of the patient visit.

Katanin maintains meiotic metaphase chromosome alignment and spindle structure in vivo and has multiple effects on microtubules in vitro.

Assembly of Caenorhabditis elegans female meiotic spindles requires both MEI-1 and MEI-2 subunits of the microtubule-severing ATPase katanin. Strong loss-of-function mutants assemble apolar intersecting microtubule arrays, whereas weaker mutants assemble bipolar meiotic spindles that are longer than wild type. To determine whether katanin is also required for spindle maintenance, we monitored metaphase I spindles after a fast-acting mei-1(ts) mutant was shifted to a nonpermissive temperature. Within 4 min of temperature shift, bivalents moved off the metaphase plate, and microtubule bundles within the spindle lengthened and developed a high degree of curvature. Spindles eventually lost bipolar structure. Immunofluorescence of embryos fixed at increasing temperature indicated that MEI-1 was lost from spindle microtubules before loss of ASPM-1, indicating that MEI-1 and ASPM-1 act independently at spindle poles. We quantified the microtubule-severing activity of purified MEI-1/MEI-2 complexes corresponding to six different point mutations and found a linear relationship between microtubule disassembly rate and meiotic spindle length. Previous work showed that katanin is required for severing at points where two microtubules intersect in vivo. We show that purified MEI-1/MEI-2 complexes preferentially sever at intersections between two microtubules and directly bundle microtubules in vitro. These activities could promote parallel/antiparallel microtubule organization in meiotic spindles.

Length of stay by route of contrast administration for diagnosis of appendicitis by computed-tomography scan.

OBJECTIVES: Studies show equivalent accuracy of abdominal computed tomography (CT) using rectal contrast compared with oral contrast for diagnosing appendicitis. The authors evaluated whether emergency department (ED) length of stay and satisfaction differed by route of contrast administration for abdominal CT. METHODS: This before-and-after intervention evaluated adult patients in an urban academic ED who were undergoing abdominal CT to screen for suspected appendicitis. Phase 1 subjects had a CT after oral contrast. Phase 2 patients had a CT after rectal contrast infused by gravity drip. Patients were interviewed after CT scan to assess satisfaction and discomfort. The primary outcome was ED length of stay. Medians, 95% binomial confidence intervals (CI), and Wilcoxon rank sum test of differences were calculated. RESULTS: One hundred twelve patients were enrolled; half received rectal contrast. There was a significant decrease in length of stay for patients who were administered rectal contrast (261 min, 95% CI = 236 to 305 min) vs. oral contrast (332 min, 95% CI = 299 to 362 min), p = 0.009. Although subjects in the rectal-contrast group waited 65 minutes longer than did oral-contrast patients before receiving contrast after the CT order, the time from contrast administration to CT was 13 minutes, vs. 150 minutes for patients receiving oral contrast (p < 0.001). Patient satisfaction and discomfort did not differ by route of contrast administration. CONCLUSIONS: Rectal contrast for patients undergoing abdominal CT to rule out appendicitis reduced ED length of stay by more than an hour and did not affect patient satisfaction or discomfort. Rectal-contrast administration for abdominal CT may significantly shorten patient throughput time for individuals undergoing evaluation for appendicitis.