Apolar peptide models for conformational heterogeneity, hydration, and packing of polypeptide helices: crystal structure of hepta- and octapeptides containing α-aminoisobutyric acid

The crystal structures of two helical peptides Boc-Val-Ala-Leu-Aib-Val-Ala-Leu-OMe (VALU-7) and Boc-Val-Ala-Leu-Aib-Val-Ala-Leu-Aib-OMe (VALU-8) have been determined to a resolution of 1.0 and 0.9 Å, respectively. Both the seven and eight residue peptides crystallize with two conformers per asymmetric unit. The VALU-8 conformers are completely helical and differ only at the C-terminus by a sign reversal of the φ ψ angles of the last residue. One of the VALUE-7 conformers occurs as a normal -helix, whereas in the other, the N(7) = O(3) α-type hydrogen bond is ruptured by the entry of a water molecule (W) into the helix, which in turn makes hydrogen bonds N(7)W = 2.97 Å and O(3) = 2.77 Å. The other side of the water molecule is surrounded by a hydrophobic pocket. These two conformers give a static representation of a step in a possible helix unwinding or folding process. In the value-8 crystal the helices aggregate in a parallel mode, whereas the aggregation is antiparallel in the VALUE-7 crystal. The crystal parameters are VALUE-7 crystal. The crystal parameters are VALUE-7, P21, a = 10.203 (3) Å, b = 19.744 (6) Å, c = 22.561 (6) Å, α = 96.76°, Z = 4, C38, H69N7O10·0.5 H2O, R = 6.65% for 3674 reflections observed gt; 3σ(F): and VALU-8, P21, a; = 10.596 (4) Å, b = 27.57 (6) Å, c = 17.745 (5) Å, β = 95.76 (3)°, Z = 4, C42H76N76O11·0.25 CH3OH, R = 6.63% for 4701 reflections observed gt; 3σ(F)

Helix packing of leucine-rich peptides: A parallel leucine ladder in the structure of Boc-Aib-Leu-Aib-Aib-Leu-Leu-Leu-Aib-Leu-Aib-OMe

The packing of peptide helices in crystals of the leucine-rich decapeptide Boc-Aib-Leu-Aib-Aib-Leu-Leu-Leu-Aib-Leu-Aib-OMe provides an example of ladder-like leucylleucyl interactions between neighboring molecules. The peptide molecule forms a helix with five 5→1 hydrogen bonds and two 4→1 hydrogen bonds near the C terminus. Three head-to-tail NH c O = C hydrogen bonds between helices form continuous columns of helices in the crystal. The helicial columns associate in an antiparallel fashion, except for the association of Leu Leu side chains, which occurs along the diagonal of the cell where the peptide helices are parallel. The peptide, with formula C56H102N10O13, crystallizes in space group P212121 with Z = 4 and cell parameters a = 16.774(3) Å, b = 20.032(3) Å and c = 20.117(3) Å; overall agreement factor R = 10.7% for 2014 data with |Fobs| <3σ(F); resolution 1.0 Å

Conformation of a 16-residue zervamicin IIA analog peptide containing three different structural features: 3<SUB>10</SUB>-helix, &#945;-helix, and &#946;-bend ribbon

Boc-Trp-Ile-Ala-Aib-Ile-Val-Aib-Leu-Aib- Pro-Ala-Aib-Pro-Aib-Pro-Phe-OMe (where Boc is t-butoxycarbonyl and Aib is a-aminoisobutyric acid), a synthetic apolar analog of the membrane-active fungal peptide antibiotic zervamycin IIA, crystallizes in space group P1 with Z = 1 and cell parameters a = 9.086 &#177; 0.002 A:, b = 10.410 &#177; 0.002 A, c = 28.188 &#177; 0.004 A, &#945; = 86.13 &#177; 0.01&#176;, &#946; = 87.90 &#177; 0.01&#176;, and &#947; = 89.27 &#177; 0.01&#176;; overall agreement factor R = 7.3% for 7180 data (Fo &gt; 3&#963;) and 0.91-A resolution. The peptide backbone makes a continuous spiral that begins as a 310-helix at the N-terminus, changes to an &#945;-helix for two turns, and ends in a spiral of three &#946;-bends in a ribbon. Each of the &#946;-bends contains a proline residue at one of the corners. The torsion angles &#966;i range from -51&#176; to -91&#176; (average value -64&#176;), and the torsion angles &#968;i range from -1&#176; to -46&#176; (average value -31&#176;). There are 10 intramolecular NH-..OC hydrogen bonds in the helix and two direct head-to-tail hydrogen bonds between successive molecules. Two H20 and two CH3OH solvent molecules fill additional space with appropriate hydrogen bonding in the head-to-tail region, and two additional H20 molecules form hydrogen bonds with carbonyl oxygens near the curve in the helix at Pro-10. Since there is only one peptide molecule per cell in space group P1, the molecules repeat only by translation, and consequently the helices pack parallel to each other

Aqueous channels within apolar peptide aggregates: solvated helix of the &#945;-aminoisobutyric acid (Aib)-containing peptide Boc-(Aib-Ala-Leu)<SUB>3</SUB>-Aib-OMe.2H<SUB>2</SUB>O.CH<SUB>3</SUB>OH in crystals

Although the peptide Boc-Aibl-Ala2-Leu3- Aib4-Ala5-Leu6-Aib7-Ala8-Leu9-Aib10-OMe [with a t-butoxycarbonyl (Boc) blocking group at the amino terminus, a methyl ester (OMe) at the carboxyl terminus, and four &#945;-aminoisobutyric (Aib) residues] has a 3-fold repeat of residues, the helix formed by the peptide backbone is irregular. The carboxyl-terminal half assumes an &#945;-helical form with torsion angles ) and &#968; of approximately -60&#176; and -45&#176;, respectively, whereas the amino-terminal half is distorted by an insertion of a water molecule between the amide nitrogen of Ala5 [N(5)] and the carbonyl oxygen of Ala2 [0(2)]. The water molecule W(1) acts as a bridge by forming hydrogen bonds N(5).W(1) (2.93 A) and W(1)---0(2) (2.86 A). The distortion of the helix exposes the carbonyl oxygens of Aib1 and Aib4 to the outside environment, with the consequence that the helix assumes an amphiphilic character despite having all apolar residues. Neighboring helices in the crystal run in antiparallel directions. On one side of a helix there are only hydrophobic contacts with efficient interdigitation of leucine side chains with those from the neighboring helix. On the other side of the helix there are hydrogen bonds between protruding carbonyl oxygens and four water molecules that separate two neighboring helices. Along the helix axis the helices bind head-to-tail with a direct hydrogen bond N(2)-0(9) (3.00 A). Crystals grown from methanol/water solution are in space group P21, with a = 15.778 &#177; 0.004 A, b = 11.228 &#177; 0.002 A, c = 18.415 &#177; 0.003 A, &#946; = 102.10 &#177; 0.02&#176; and two formula units per cell for C49H88N1003. 2H2O.CH3OH. The overall agreement factor R is 7.5% for 3394 reflections observed with intensities &gt; 3&#963;(F), and the resolution is 0.90 A

Nursing Roles and Strategies in End-of-Life Decision Making in Acute Care: A Systematic Review of the Literature

The objective of this paper is to analyze the literature concerning nurses' roles and strategies in EOL decision making in acute care environments, synthesize the findings, and identify implications for future research. We conducted searches in CINAHL and PubMed, using a broad range of terms. The 44 articles retained for review had quantitative and qualitative designs and represented ten countries. These articles were entered into a matrix to facilitate examining patterns, themes, and relationships across studies. Three nursing roles emerged from the synthesis of the literature: information broker, supporter, and advocate, each with a set of strategies nurses use to enact the roles. Empirical evidence linking these nursing roles and strategies to patients and family members outcomes is lacking. Understanding how these strategies and activities are effective in helping patients and families make EOL decisions is an area for future research

Improving the accuracy and utility of harmful algal bloom forecasting systems

Author Posting. © The Author(s), 2006. This is the author's version of the work. It is posted here by permission of Geological Society for the Micropalaeontological Society for personal use, not for redistribution. The definitive version was published in Biological and Geological Perspectives of Dinoflagellates, edited by Jane Lewis, F. Marret, L.Bradley, :141-147. London: Geological Society for the Micropalaeontological Society, 2013. ISBN: 9781862393684.One of the goals of harmful algal bloom (HAB) research has been to develop predictive capabilities for blooms. Major steps have been made towards this goal, including the development of physical-biological models of HAB species that simulate bloom dynamics in specific regions. In the Gulf of Maine region of the northeastern U.S., models have been developed that have considerable skill in simulating blooms of Alexandrium fundyense, the causative organism for paralytic shellfish poisoning (PSP) outbreaks in the region. This model is now being used for both short-term and long-term forecasts. This paper describes several ongoing activities that will improve the accuracy and usefulness of the model and forecasts. These include efforts to streamline or minimize the sampling and analysis requirements of annual A. fundyense cyst surveys, efforts to quantitatively describe or characterize the severity of predicted outbreaks, and plans to obtain real-time data on Alexandrium cell abundance and toxicity that can be assimilated into the models. Together, these and other activities are moving us towards an operational forecasting system for Alexandrium blooms in the region.Woods Hole Center for Oceans and Human Health, National Science Foundation (NSF) Grants OCE-0430724, and OCE-0911031; and National Institute of Environmental Health Sciences (NIEHS) Grant 1-P50-ES012742-01, the ECOHAB Grant program through NOAA Grants NA06NOS4780245 and NA09NOS4780193, and the MERHAB Grant program through NOAA Grant NA11NOS4780025

Novel Microdialysis Technique Reveals a Dramatic Shift in Metabolite Secretion during the Early Stages of the Interaction between the Ectomycorrhizal Fungus Pisolithus microcarpus and Its Host Eucalyptus grandis

The colonisation of tree roots by ectomycorrhizal (ECM) fungi is the result of numerous signalling exchanges between organisms, many of which occur before physical contact. However, information is lacking about these exchanges and the compounds that are secreted by each organism before contact. This is in part due to a lack of low disturbance sampling methods with sufficient temporal and spatial resolution to capture these exchanges. Using a novel in situ microdialysis approach, we sampled metabolites released from Eucalyptus grandis and Pisolithus microcarpus independently and during indirect contact over a 48-h time-course using UPLC-MS. A total of 560 and 1530 molecular features (MFs; ESI- and ESI+ respectively) were identified with significant differential abundance from control treatments. We observed that indirect contact between organisms altered the secretion of MFs to produce a distinct metabolomic profile compared to either organism independently. Many of these MFs were produced within the first hour of contact and included several phenylpropanoids, fatty acids and organic acids. These findings show that the secreted metabolome, particularly of the ECM fungus, can rapidly shift during the early stages of pre-symbiotic contact and highlight the importance of observing these early interactions in greater detail. We present microdialysis as a useful tool for examining plant-fungal signalling with high temporal resolution and with minimal experimental disturbance

Aggregation studies in crystals of apolar helical peptides: Boc-Aib-Val-Ala-Leu-Aib-Val-Ala-Leu-Aib-OMe

In the crystal, the backbone of Boc-(Aib-Val-Ala-Leu)2-Aib-OMc adopts a helical form with four &#945;-type hydrogen bonds in the middle, flanked by 310-type hydrogen bonds at either end. The helical molecules stack in columns with head-to-tail hydrogen bonds, either directly between NH and CO, or bridged by solvent molecules. The packing of the helices is parallel, even in space group P21. Cell parameters are a = 9.837(2) &#197;. b = 15.565(3) &#197;. c = 20.087(5) &#197;, &#946;= 96.42(2)&#176;. dcalc= 1.091 g/cm3 for C46H83,N9O12&#183; 1.5H2O&#183;0.67CH3OH. There appears to be some hydration of the backbone in this apolar helix

EZH2 or HDAC1 Inhibition Reverses Multiple Myeloma-Induced Epigenetic Suppression of Osteoblast Differentiation

In multiple myeloma, osteolytic lesions rarely heal because of persistent suppressed osteoblast differentiation resulting in a high fracture risk. Herein, chromatin immunoprecipitation analyses reveal that multiple myeloma cells induce repressive epigenetic histone changes at the Runx2 locus that prevent osteoblast differentiation. The most pronounced multiple myeloma-induced changes were at the Runx2-P1 promoter, converting it from a poised bivalent state to a repressed state. Previously, it was observed that multiple myeloma induces the transcription repressor GFI1 in osteoblast precursors, which correlates with decreased Runx2 expression, thus prompting detailed characterization of the multiple myeloma and TNFα-dependent GFI1 response element within the Runx2-P1 promoter. Further analyses reveal that multiple myeloma-induced GFI1 binding to Runx2 in osteoblast precursors and recruitment of the histone modifiers HDAC1, LSD1, and EZH2 is required to establish and maintain Runx2 repression in osteogenic conditions. These GFI1-mediated repressive chromatin changes persist even after removal of multiple myeloma. Ectopic GFI1 is sufficient to bind to Runx2, recruit HDAC1 and EZH2, increase H3K27me3 on the gene, and prevent osteogenic induction of endogenous Runx2 expression. Gfi1 knockdown in MC4 cells blocked multiple myeloma-induced recruitment of HDAC1 and EZH2 to Runx2, acquisition of repressive chromatin architecture, and suppression of osteoblast differentiation. Importantly, inhibition of EZH2 or HDAC1 activity in pre-osteoblasts after multiple myeloma exposure in vitro or in osteoblast precursors from patients with multiple myeloma reversed the repressive chromatin architecture at Runx2 and rescued osteoblast differentiation.Implications: This study suggests that therapeutically targeting EZH2 or HDAC1 activity may reverse the profound multiple myeloma-induced osteoblast suppression and allow repair of the lytic lesions

Validation and Generalizability of Preoperative PROMIS Scores to Predict Postoperative Success in Foot and Ankle Patients

Background: A recent publication reported preoperative Patient-Reported Outcomes Measurement Instrumentation System (PROMIS) scores to be highly predictive in identifying patients who would and would not benefit from foot and ankle surgery. Their applicability to other patient populations is unknown. The aim of this study was to assess the validation and generalizability of previously published preoperative PROMIS physical function (PF) and pain interference (PI) threshold t scores as predictors of postoperative clinically meaningful improvement in foot and ankle patients from a geographically unique patient population. Methods: Prospective PROMIS PF and PI scores of consecutive patient visits to a tertiary foot and ankle clinic were obtained between January 2014 and November 2016. Patients undergoing elective foot and ankle surgery were identified and PROMIS values obtained at initial and follow-up visits (average, 7.9 months). Analysis of variance was used to assess differences in PROMIS scores before and after surgery. The distributive method was used to estimate a minimal clinically important difference (MCID). Receiver operating characteristic curve analysis was used to determine thresholds for achieving and failing to achieve MCID. To assess the validity and generalizability of these threshold values, they were compared with previously published threshold values for accuracy using likelihood ratios and pre- and posttest probabilities, and the percentages of patients identified as achieving and failing to achieve MCID were evaluated using χ2 analysis. Results: There were significant improvements in PF (P \u3c .001) and PI (P \u3c .001) after surgery. The area under the curve for PF (0.77) was significant (P \u3c .01), and the thresholds for achieving MCID and not achieving MCID were similar to those in the prior study. A significant proportion of patients (88.9%) identified as not likely to achieve MCID failed to achieve MCID (P = .03). A significant proportion of patients (84.2%) identified as likely to achieve MCID did achieve MCID (P \u3c .01). The area under the curve for PROMIS PI was not significant. Conclusions: PROMIS PF threshold scores from published data were successful in classifying patients from a different patient and geographic population who would improve with surgery. If functional improvement is the goal, these thresholds could be used to help identify patients who will benefit from surgery and, most important, those who will not, adding value to foot and ankle health care. Level of evidence: Level II, Prospective Comparative Stud