Selective peptidic and peptidomimetic inhibitors of Candida albicans myristoylCoA: protein N-myristoyltransferase: a new approach to antifungal therapy

MyristoylCoA: protein N-myristoyltransferase (NMT) catalyzes the cotranslational covalent attachment of a rare cellular fatty acid, myristate, to the N-terminal Gly residue of a variety of eukaryotic proteins. The myristoyl moiety is often essential for expression of the biological functions for these proteins. Attachment of C14:0 alone provides barely enough hydrophobicity to allow stable association with membranes. The partitioning of N-myrisotylproteins is therefore often modulated by "switches" that function through additional covalent or noncovalent modifications. Candida albicans, the principal cause of systemic fungal infection in immunocompromised humans, contains a single NMT gene that is essential for its viability. The functional properties of the acylCoA binding site of human and C. albicans NMT are very similar. However, there are distinct differences in their peptide binding sites. An ADP ribosylation factor (Arf) is included among the few cellular protein substrates of the fungal enzyme. Alanine scanning mutagenesis of an octapeptide derived from an N-terminal Arf sequence (GLYASKLS-NH2) disclosed that Gly1, Ser5, and Lys6 play predominant roles in binding. ALYASKLS-NH2 is an inhibitor competitive for peptide [Ki(app) = 15.3 +/- 6.4 microM] and noncompetitive for myristoylCoA. Remarkably, replacement of the N-terminal tetrapeptide with an 11-aminoundecanoyl group results in a competitive inhibitor (11-aminoundecanoyl-SKLS-NH2) that is approximately 40-fold more potent [Ki(app) = 0.40 +/- 0.03 microM] than the starting octapeptide. Removal of Leu-Ser from the C-terminus generates a competitive dipeptide inhibitor (11-aminoundecanoyl-SK-NH2) with a Ki(app) of 11.7 +/- 0.4 microM, equivalent to that of the starting octapeptide. A derivative dipeptide inhibitor containing a C-terminal N-cyclohexylethyl lysinamide moiety has the advantage of being more potent (IC50 = 0.11 +/- 0.03 microM) and resistant to digestion by cellular carboxypeptidases. Rigidifying the flexible aminoundecanoyl chain results in very potent general NMT inhibitors (IC50 = 40-50 nM). Substituting a 2-methylimidazole for the N-terminal amine and adding a benzylic alpha-methyl group with R stereochemistry to the rigidifying element produces even more potent inhibitors (IC50 = 20-50 nM) that are up to 500-fold selective for the fungal compared to human enzyme. A related less potent member of this series of compounds is fungistatic. Its growth inhibitory effects are associated with a reduction in cellular protein N-myristoylation, judged using cellular Arf as a reporter. These studies establish that NMT is a new antifungal target.     

A framework to practical predictive maintenance modeling for multi-state systems

A simple practical framework for predictive maintenance (PdM)-based scheduling of multi-state systems (MSS) is developed. The maintenance schedules are derived from a system-perspective using the failure times of the overall system as estimated from its performance degradation trends.

The system analyzed in this work is a flow transmission water pipe system. The various factors influencing PdM-based scheduling are identified and their impact on the system reliability and performance are quantitatively studied. The estimated times to replacement of the MSS may also be derived from the developed model.

The results of the model simulation demonstrate the significant impact of maintenance quality and the criteria for the call for maintenance (user demand) on the system reliability and mean performance characteristics. A slight improvement in maintenance quality is found to postpone the system replacement time by manifold. The consistency in the quality of maintenance work with minimal variance is also identified as a very important factor that enhances the system's future operational and downtime event predictability.

The studies also reveal that in order to reduce the frequency of maintenance actions, it is necessary to lower the minimum user demand from the system if possible, ensuring at the same time that the system still performs its intended function effectively.

The model proposed can be utilized to implement a PdM program in the industry with a few modifications to suit the individual industrial systems’ needs.     

Multimodal social intelligence in a real-time dashboard system

Social Networks provide one of the most rapidly evolving data sets in existence today. Traditional Business Intelligence applications struggle to take advantage of such data sets in a timely manner. The BBC SoundIndex, developed by the authors and others, enabled real-time analytics of music popularity using data from a variety of Social Networks. We present this system as a grounding example of how to overcome the challenges of working with this data from social networks. We discuss a variety of technologies to implement near real-time data analytics to transform Social Intelligence into Business Intelligence and evaluate their effectiveness in the music domain. The SoundIndex project helped to highlight a number of key research areas, including named entity recognition and sentiment analysis in Informal English. It also drew attention to the importance of metadata aggregation in multimodal environments. We explored challenges such as drawing data from a wide set of sources spanning a myriad of modalities, developing adjudication techniques to harmonize inputs, and performing deep analytics on extremely challenging Informal English snippets. Ultimately, we seek to provide guidance on developing applications in a variety of domains that allow an analyst to rapidly grasp the evolution in the social landscape, and show how to validate such a system for a real-world application.     

Position of helical kinks in membrane protein crystal structures and the accuracy of computational prediction

The structural features of helical transmembrane (TM) proteins, such as helical kinks, tilts, and rotational orientations are important in modulation of their function and these structural features give rise to functional diversity in membrane proteins with similar topology. In particular, the helical kinks caused by breaking of the backbone hydrogen bonds lead to hinge bending flexibility in these helices. Therefore it is important to understand the nature of the helical kinks and to be able to reproduce these kinks in structural models of membrane proteins. We have analyzed the position and extent of helical kinks in the transmembrane helices of all the crystal structures of membrane proteins taken from the MPtopo database, which are about 405 individual helices of length between 19 and 35 residues. 44% of the crystal structures of TM helices showed a significant helical kink, and 35% of these kinks are caused by prolines. Many of the non-proline helical kinks are caused by other residues like Ser and Gly that are located at the center of helical kinks. The side chain of Ser makes a hydrogen bond with the main chain carbonyl of the i − 4th or i + 4th residue thus making a kink. We have also studied how well molecular dynamics (MD) simulations on isolated helices can reproduce the position of the helical kinks in TM helices. Such a method is useful for structure prediction of membrane proteins. We performed MD simulations, starting from a canonical helix for the 405 TM helices. 1 ns of MD simulation results show that we can reproduce about 79% of the proline kinks, only 59% of the vestigial proline kinks and 18% of the non-proline helical kinks. We found that similar results can be obtained from choosing the lowest potential energy structure from the MD simulation. 4–14% more of the vestigial prolines were reproduced by replacing them with prolines before performing MD simulations, and changing the amino acid back to proline after the MD simulations. From these results we conclude that the position of the helical kinks is dependent on the TM sequence. However the extent of helical kinking may depend on the packing of the rest of the protein and the lipid bilayer.     

Benchmarking the perioperative process: III. Effects of regional anesthesia clinical pathway techniques on process efficiency and recovery profiles in ambulatory orthopedic surgery

STUDY OBJECTIVES: (1) To incorporate regional anesthesia options for common outpatient orthopedic surgery into clinical pathways; (2) to use the clinical pathway format and the Procedural Times Glossary published by the Association of Anesthesia Clinical Directors (AACD) as management tools to measure postoperative same-day surgery processes and discharge outcomes; and (3) to determine the effects of general, regional, and combined general-regional anesthesia on these processes and outcomes. DESIGN: Hospital database and patient chart review of consecutive patients undergoing anterior cruciate ligament reconstruction (ACLR) during academic years (AY) 1995-1996 and 1996-1997. Patient data from AY 1995-1996, during which no intraoperative anesthesia clinical pathway existed, served as historical controls. Data from AY 1996-1997, during which intraoperative anesthesia clinical pathways were used, served as the treatment group. SETTING: Ambulatory surgery center in a teaching hospital. MEASUREMENTS AND MAIN RESULTS: The records of 503 ASA physical status I and II patients were reviewed. 1996-1997 patients selected general anesthesia (+/- femoral nerve block) or epidural anesthesia, after which the remainder of the perioperative anesthesia process was standardized with respect to the drugs and equipment used. 1995-1996 patients did not necessarily have a choice in anesthesia technique and did not have a standardized perioperative anesthetic course with respect to specific drugs and supplies. Intervals described in the AACD Procedural Times Glossary, anesthesia drug and supply costs, and patient outcome variables (postoperative nursing interventions required and unexpected admissions), as influenced by anesthesia technique used, were measured. Combined general-regional anesthesia care for ACLR in 1996-1997, when compared with general anesthesia alone, led to increased pharmacy and materials costs and increased turnover time. However, patients with the combined technique showed improved recovery profiles and lower unexpected admission rates, and they required fewer nursing interventions for common postoperative symptoms. Patients receiving epidural anesthesia showed discharge outcomes similar to those patients receiving general anesthesia with femoral nerve block. Postanesthesia care unit bypass (fast-tracking) was more likely in clinical pathway regional anesthesia patients, when compared with the clinical pathway general anesthesia used. CONCLUSIONS: Clinical pathway regional anesthesia care for outpatient orthopedics may have a significant role in simultaneously containing costs and improving both process efficiency and patient outcomes.