Medicine and Health Sciences Commons^™

Structural And Thermodynamic Basis For The Binding Of Tmc114, A Next-Generation Human Immunodeficiency Virus Type 1 Protease Inhibitor, Nancy King, Moses Prabu-Jeyabalan, Ellen Nalivaika, Piet Wigerinck, Marie-Pierre De Bethune, Celia Schiffer

Celia A. Schiffer

TMC114, a newly designed human immunodeficiency virus type 1 (HIV-1) protease inhibitor, is extremely potent against both wild-type (wt) and multidrug-resistant (MDR) viruses in vitro as well as in vivo. Although chemically similar to amprenavir (APV), the potency of TMC114 is substantially greater. To examine the basis for this potency, we solved crystal structures of TMC114 complexed with wt HIV-1 protease and TMC114 and APV complexed with an MDR (L63P, V82T, and I84V) protease variant. In addition, we determined the corresponding binding thermodynamics by isothermal titration calorimetry. TMC114 binds approximately 2 orders of magnitude more tightly to the wt enzyme …

Lack Of Synergy For Inhibitors Targeting A Multi-Drug-Resistant Hiv-1 Protease, Nancy King, Laurence Melnick, Moses Prabu-Jeyabalan, Ellen Nalivaika, Shiow-Shong Yang, Yun Gao, Xiaoyi Nie, Charles Zepp, Donald Heefner, Celia Schiffer

Celia A. Schiffer

The three-dimensional structures of indinavir and three newly synthesized indinavir analogs in complex with a multi-drug-resistant variant (L63P, V82T, I84V) of HIV-1 protease were determined to approximately 2.2 A resolution. Two of the three analogs have only a single modification of indinavir, and their binding affinities to the variant HIV-1 protease are enhanced over that of indinavir. However, when both modifications were combined into a single compound, the binding affinity to the protease variant was reduced. On close examination, the structural rearrangements in the protease that occur in the tightest binding inhibitor complex are mutually exclusive with the structural rearrangements …

Hydrophobic Sliding: A Possible Mechanism For Drug Resistance In Human Immunodeficiency Virus Type 1 Protease, Jennifer Foulkes-Murzycki, Walter Scott, Celia Schiffer

Celia A. Schiffer

Hydrophobic residues outside the active site of HIV-1 protease frequently mutate in patients undergoing protease inhibitor therapy; however, the mechanism by which these mutations confer drug resistance is not understood. From analysis of molecular dynamics simulations, 19 core hydrophobic residues appear to facilitate the conformational changes that occur in HIV-1 protease. The hydrophobic core residues slide by each other, exchanging one hydrophobic van der Waal contact for another, with little energy penalty, while maintaining many structurally important hydrogen bonds. Such hydrophobic sliding may represent a general mechanism by which proteins undergo conformational changes. Mutation of these residues in HIV-1 protease …

Viability Of A Drug-Resistant Human Immunodeficiency Virus Type 1 Protease Variant: Structural Insights For Better Antiviral Therapy, Moses Prabu-Jeyabalan, Ellen A. Nalivaika, Nancy M. King, Celia A. Schiffer

Celia A. Schiffer

Under the selective pressure of protease inhibitor therapy, patients infected with human immunodeficiency virus (HIV) often develop drug-resistant HIV strains. One of the first drug-resistant mutations to arise in the protease, particularly in patients receiving indinavir or ritonavir treatment, is V82A, which compromises the binding of these and other inhibitors but allows the virus to remain viable. To probe this drug resistance, we solved the crystal structures of three natural substrates and two commercial drugs in complex with an inactive drug-resistant mutant (D25N/V82A) HIV-1 protease. Through structural analysis and comparison of the protein-ligand interactions, we found that Val82 interacts more …

Role Of Invariant Thr80 In Human Immunodeficiency Virus Type 1 Protease Structure, Function, And Viral Infectivity, Jennifer E. Foulkes-Murzycki, Moses Prabu-Jeyabalan, Deyna Cooper, Gavin J. Henderson, Janera Harris, Ronald I. Swanstrom, Celia A. Schiffer

Celia A. Schiffer

Sequence variability associated with human immunodeficiency virus type 1 (HIV-1) is useful for inferring structural and/or functional constraints at specific residues within the viral protease. Positions that are invariant even in the presence of drug selection define critically important residues for protease function. While the importance of conserved active-site residues is easily understood, the role of other invariant residues is not. This work focuses on invariant Thr80 at the apex of the P1 loop of HIV-1, HIV-2, and simian immunodeficiency virus protease. In a previous study, we postulated, on the basis of a molecular dynamics simulation of the unliganded protease, …

Contribution Of Ser386 And Ser396 To Activation Of Interferon Regulatory Factor 3, Weijun Chen, Hema Srinath, Suvana Lam, Celia Schiffer, William Royer, Kai Lin

Celia A. Schiffer

IRF-3, a member of the interferon regulatory factor (IRF) family of transcription factors, functions in innate immune defense against viral infection. Upon infection, host cell IRF-3 is activated by phosphorylation at its seven C-terminal Ser/Thr residues: (385)SSLENTVDLHISNSHPLSLTS(405). This phosphoactivation triggers IRF-3 to react with the coactivators, CREB-binding protein (CBP)/p300, to form a complex that activates target genes in the nucleus. However, the role of each phosphorylation site for IRF-3 phosphoactivation remains unresolved. To address this issue, all seven Ser/Thr potential phosphorylation sites were screened by mutational studies, size-exclusion chromatography, and isothermal titration calorimetry. Using purified proteins, we show that CBP …

Competition Between Ski And Creb-Binding Protein For Binding To Smad Proteins In Transforming Growth Factor-Beta Signaling, Weijun Chen, Suvana Lam, Hema Srinath, Celia Schiffer, William Royer, Kai Lin

Celia A. Schiffer

The family of Smad proteins mediates transforming growth factor-beta (TGF-beta) signaling in cell growth and differentiation. Smads repress or activate TGF-beta signaling by interacting with corepressors (e.g. Ski) or coactivators (e.g. CREB-binding protein (CBP)), respectively. Specifically, Ski has been shown to interfere with the interaction between Smad3 and CBP. However, it is unclear whether Ski competes with CBP for binding to Smads and whether they can interact with Smad3 at the same binding surface on Smad3. We investigated the interactions among purified constructs of Smad, Ski, and CBP in vitro by size-exclusion chromatography, isothermal titration calorimetry, and mutational studies. Here, …

Mass Spectrometry Analysis Of Hiv-1 Vif Reveals An Increase In Ordered Structure Upon Oligomerization In Regions Necessary For Viral Infectivity, Jared Auclair, Karin Green, Shivender Shandilya, James Evans, Mohan Somasundaran, Celia Schiffer

Celia A. Schiffer

HIV-1 Vif, an accessory protein in the viral genome, performs an important role in viral pathogenesis by facilitating the degradation of APOBEC3G, an endogenous cellular inhibitor of HIV-1 replication. In this study, intrinsically disordered regions are predicted in HIV-1 Vif using sequence-based algorithms. Intrinsic disorder may explain why traditional structure determination of HIV-1 Vif has been elusive, making structure-based drug design impossible. To characterize HIV-1 Vif's structural topology and to map the domains involved in oligomerization we used chemical cross-linking, proteolysis, and mass spectrometry. Cross-linking showed evidence of monomer, dimer, and trimer species via denaturing gel analysis and an additional …

Exploring The Role Of The Solvent In The Denaturation Of A Protein: A Molecular Dynamics Study Of The Dna Binding Domain Of The 434 Repressor, Celia Schiffer, Volker Dötsch, Kurt Wuthrich, Wilfred Van Gunsteren

Celia A. Schiffer

Molecular dynamics simulations of the DNA binding domain of 434 repressor are presented which aim at unraveling the role of solvent in protein denaturation. Four altered solvent models, each mimicking various possible aspects of the addition of a denaturant to the aqueous solvent, were used in the simulations to analyze their effects on the stability of the protein. The solvent was altered by selectively changing the Coulombic interaction between water and protein atoms and between different water molecules. The use of a modified solvent model has the advantage of mimicking the presence of denaturant without having denaturant molecules present in …

Structural Analysis Of Human Immunodeficiency Virus Type 1 Crf01_Ae Protease In Complex With The Substrate P1-P6., Rajintha Bandaranayake, Moses Prabu-Jeyabalan, Junko Kakizawa, Wataru Sugiura, Celia Schiffer

Celia A. Schiffer

The effect of amino acid variability between human immunodeficiency virus type 1 (HIV-1) clades on structure and the emergence of resistance mutations in HIV-1 protease has become an area of significant interest in recent years. We determined the first crystal structure of the HIV-1 CRF01_AE protease in complex with the p1-p6 substrate to a resolution of 2.8 A. Hydrogen bonding between the flap hinge and the protease core regions shows significant structural rearrangements in CRF01_AE protease compared to the clade B protease structure.

Phylogenomics Of Reichenowia Parasitica, An Alphaproteobacterial Endosymbiont Of The Freshwater Leech Placobdella Parasitica, Sebastian Kvist, Apurva Narechania, Alejandro Oceguera-Figueroa, Bella Fuks, Mark E. Siddall

Publications and Research

Although several commensal alphaproteobacteria form close relationships with plant hosts where they aid in (e.g.,) nitrogen fixation and nodulation, only a few inhabit animal hosts. Among these, Reichenowia picta, R. ornata and R. parasitica, are currently the only known mutualistic, alphaproteobacterial endosymbionts to inhabit leeches. These bacteria are harbored in the epithelial cells of the mycetomal structures of their freshwater leech hosts, Placobdella spp., and these structures have no other obvious function than housing bacterial symbionts. However, the function of the bacterial symbionts has remained unclear. Here, we focused both on exploring the genomic makeup of R. parasitica and on …

Perceptions Of Flatulence From Bean Consumption Among Adults In 3 Feeding Studies, Donna M. Winham, Andrea M. Hutchins

Donna Winham

Background: Many consumers avoid eating beans because they believe legume consumption will cause excessive intestinal gas or flatulence. An increasing body of research and the 2010 Dietary Guidelines for Americans supports the benefits of a plant-based diet, and legumes specifically, in the reduction of chronic disease risks. The purpose of the current research was to investigate the perception of increased flatulence and gastrointestinal discomfort among participants who consumed a ½ cup of beans daily for 8 or 12 weeks. Methods: Participants in three studies to test the effects of beans on heart disease biomarkers completed the same weekly questionnaire to …

An Appreciation Of Ralph Marvin Steinman (1943-2011), Carol L. Moberg

Steinman and the Nobel Prize

Posted with Permission.The Rockefeller University Press J. Exp. Med. Vol. 208 No. 12 2337-2342 www.jem.org/cgi/doi/10.1084/jem.20112294

An Examination Of Diet, Acculturation And Risk Factors For Heart Disease Among Jamaican Immigrants, Carol Renee Oladele

USF Tampa Graduate Theses and Dissertations

Background: The South Florida region is home to over 85,000 Jamaican immigrants. Yet, little is known about the dietary intakes and predictors of risk of disease within this immigrant group. An assessment of dietary intakes and the development of dietary intake methodologies specific to the Jamaican population was important as it permitted accurate estimation of the nutrient intakes of this immigrant population whose dietary habits are not well documented. In addition, nothing is known about the prevalence of risk factors for heart disease or factors influencing risk factors among this immigrant group. The purpose of this study was to assess …

Introducing Functional Data Analysis To Neuroimaging, And Vice Versa, Philip T. Reiss

Philip T. Reiss

No abstract provided.

Lipopolysaccharide Biosynthesis Without The Lipids: Substrate Recognition For Escherichia Coli Heptosyltransferasei, Daniel J. Czyzyk, Cassie Liu, Erika A. Taylor

Erika A. Taylor, Ph.D.

Heptosyltransferase I (HepI) is responsible for the transfer of l-glycero-d-manno-heptose to a 3-deoxy-α-D-oct-2-ulopyranosonic acid (Kdo) of the growing core region of lipopolysaccharide (LPS). The catalytic efficiency of HepI with the fully deacylated analogue of Escherichia coli HepI LipidA is 12-fold greater than with the fully acylated substrate, with a k(cat)/K(m) of 2.7 × 10(6) M(-1) s(-1), compared to a value of 2.2 × 10(5) M(-1) s(-1) for the Kdo(2)-LipidA substrate. Not only is this is the first demonstration that an LPS biosynthetic enzyme is catalytically enhanced by the absence of lipids, this result has significant implications for downstream enzymes that …

Front Of Pack Daily Intake Labelling On Australian Packaged Foods: Introduction And Use 2007-2009, P Williams, R Duncan, K De Agnoli, A Hull, A Owers, T Wang

Peter Williams

This study aimed to measure the extent of use of front-of-pack daily intake (DI) labelling across food categories in Australian supermarkets, and assess the level of compliance with the Australian Food and Grocery Council (AFGC) guidelines. Surveys of six supermarkets in the Illawarra region of New South Wales were conducted twice a year in 2007, 2008 and 2009. The number of products with DI labelling increased from 58 in February 2007 to 1939 in August 2009 and appears to be growing strongly. The greatest number of products with the labelling are in the biscuits and crackers, cooking sauces, breakfast cereals, …

Breast Cancer Detection Messages In Australian Print Media Advertising - Are They Promoting Correct Information?, Sandra C. Jones, John R. Rossiter

Sandra Jones

With breast cancer now the leading cause of cancer deaths among women in the western world, correct information about detection and treatment is vitally important. The media are often accused of conveying inaccurate information in their editorial coverage of health issues, but few studies have examined the accuracy of information in media advertising. In this study of breast cancer detection ads in Australian magazines and newspapers, many instances of misleading information were found. As the print media have a major influence on women's health beliefs, these findings have serious implications for health communication policy regarding socially responsible advertising.

Disease Awareness Advertising: Women's Intentions Following Exposure, Danika Hall, Sandra C. Jones, Donald C. Iverson

Sandra Jones

Background: In Australia, where direct to consumer advertising of prescription medicines is prohibited, pharmaceutical companies can sponsor disease awareness advertising targeting consumers. This study examined the impact of disease awareness advertising exposure on older women's reported behavioural intentions. Method: Women were approached in a shopping centre and randomly assigned mock advertisements for two health conditions. Disease information and sponsors were manipulated. Results: Two hundred and forty-one women responded to 466 advertisements. Almost half reported an intention to ask their doctor for a prescription or referral as a result of seeing the advertisement, but more reported they would talk to their …

Alcohol Energy Drinks: Is It Time For The Alcohol Industry To Demonstrate Some Corporate Social Responsibility?, Sandra Jones

Sandra Jones

There is increasing evidence of association between AEDs and both increased alcohol consumption and increased alcohol-related harm. Australia lags behind other countries in researching and regulating (alcoholic and non-alcoholic) energy drinks. Twelve focus groups were conducted with 12-17 year olds in a capital city, regional city and rural city in NSW, Australia. AEDs were popular among teenagers in urban, regional and rural areas of NSW. Adolescents liked AEDs because they provide increased energy while allowing them to feel the effects of alcohol, and because of their similarity to non-alcohol energy drinks and soft drinks. Given the industry’s stated position on …

Point-Of-Sale Alcohol Promotions In The Perth And Sydney Metropolitan Areas, Sandra C. Jones, Lance R. Barrie

Sandra Jones

There is considerable evidence that reducing the price of alcohol increases the amount consumed, particularly among young people. However, there is an absence of research on other point-of-sale (POS) alcohol marketing strategies - such as those that ‘reward’ purchasers with free gifts. The purpose of the present study was to document the nature and extent of POS alcohol promotions in bottle shops in two Australian capital cities. An audit tool was developed and point-of-sale promotions were found to be ubiquitous, with 416 promotions identified across 24 audits (an average of 17.3 promotions per outlet; 19.8 in Sydney and 14.9 in …

Methodological Difficulties In A Systematic Review Of Social Marketing For Sun Protection: Implications For Research And Practice, Sandra C. Jones, Keryn Johnson

Sandra Jones

This paper reports on a systematic review of all published skin cancer primary prevention interventions aimed at improving the sun protection of children and adolescents, from 1980 to 2005. This was undertaken to inform the development of future social marketing campaigns for the prevention of skin cancer. Rather than reporting the findings of the review, in terms of conclusions drawn about the effectiveness of interventions, this paper focuses on the systematic review process itself - identifying and discussing the methodological difficulties that arose in conducting this review. These difficulties, from lack of information on the development and theoretical background of …

Evaluating The Impact Of The Dandenong Public Drinking Campaign, Lance R. Barrie, Sandra C. Jones, Dona Macik, Peter Griffith

Sandra Jones

In 2009, Dandenong launch a campaign targeting drinking in public places using stencils, posters and stickers in community settings and bottle-shops. An evaluation was undertaken and 152 participants completed a survey where elements of the Health Belief Model (HBM) were explored. Over 65% of respondents reported that they had seen at least 4 different advertisements, suggesting that the campaign dissemination was very effective. The campaign had a high recall rate and over 25% of people had either stopped or intended to stop drinking in public places. Overall, awareness of the negative effects of drinking in public increased; however no follow …

Process Evaluation Of An Innovative Sun Protection Intervention Targeting Adolescents, Sandra C. Jones, Keryn M. Johnson, Donald C. Iverson, Jeffrey Thom

Sandra Jones

The purpose of the current intervention was to target a defined segment of the adolescent population with a sun protection ‘offering’ that positioned sun protection as beneficial and addressed identified barriers (particularly inconvenience and image). A community intervention was conducted in one defined geographic region over the 2009/2010 Summer school holidays. Key elements of the intervention included the distribution of augmented products, promotional materials with a pre-tested impactful message, and partnerships with community and commercial organisations. The intervention was successful in creating interest and attention among adolescents. This paper reports on the process evaluations, focusing on barriers, facilitators and lessons …

"At Least I'M Not Drink-Driving": Formative Research For A Social Marketing Campaign To Reduce Drug-Driving Among Young Drivers, Lance R. Barrie, Sandra C. Jones, Elizabeth Wiese

Sandra Jones

This paper reports on a qualitative study designed to examine young drivers’ knowledge and attitudes regarding drug-driving, as the formative research for a potential drug-driving social marketing program in the Australian Capital Territory (ACT). Drug driving has been found to be associated with motor vehicle accidents, particularly among younger drivers. However, the potential for social marketing in this area has received little attention. This study found that young people were not aware of the effects of drugs on driving, formed their perceptions of risk (both of getting caught and of impaired driving) based on other people’s experiences, and felt that …

General Practice Registrars: Attitudes Of Older Patients, Andrew D. Bonney, Sandra C. Jones, Lyn Phillipson, Donald C. Iverson

Sandra Jones

Previous research indicates that older patients may be less willing to consult general practice registrars (GPRs), reducing training opportunities in chronic/complex care. This survey explores older patients’ attitudes in order to inform models of interaction that would be acceptable to patients. METHODS Ten training general practices distributed questionnaires for self completion to 50 patients aged 60 years and over. Chi-square, Spearman’s rho and logistic regression were used for analysis. RESULTS The response rate was 47%. Ninetysix percent wanted ongoing contact with their general practitioner if they saw a GPR. Twenty-four percent were comfortable with GPR chronic/complex care, increasing to 73% …

Case Study In Academic And Industry Collaboration: The Development Of An Adolescent Targeted Sun Protection Intervention In Nsw, Melinda Williams, Sandra Jones, Donald Iverson, Peter Caputi, Sofia Potente

Sandra Jones

Academic and industry collaboration is increasingly identified as a critical element in the future health of Australians through linking theory and practice, with the major priority for academic institutions being the identification of new knowledge and the transfer of this knowledge into changes in policy and health services. Collaborations between academia and industry are increasingly encouraged in Australia by research funding schemes such as ARCLinkage and, more recently, NHMRC Partnerships. While a recent US study suggests that such schemes have a moderate effect on academics’ propensity to work with industry (Bozeman and Gaughan, 2007), industry groups have recognised the value …

Preparing For An Ageing Population: A Survey Of Older Patients' Attitudes To General Practice Registrars, Andrew D. Bonney, Sandra C. Jones, Lyn Phillipson, Donald C. Iverson

Sandra Jones

The ageing population makes it imperative to provide appropriate training for general practice registrars (GPRs) in the community-based care of older patients. However, data suggest that older patients may be less willing to consult GPRs for chronic/complex care; adversely affecting training opportunities and potentially the satisfaction of older patients in training practices. This cross-sectional study was undertaken to investigate this concern in the Australian context and develop models of older patient-GPR interaction that are acceptable to patients.

The Effect Of Exercise On The Skeletal Muscle Phospholipidome Of Rats Fed A High-Fat Diet, Todd W. Mitchell, N. Turner, Paul Else, Anthony J. Hulbert, John Hawley, Jong Sam Lee, Clinton Bruce, Stephen J. Blanksby

Stephen Blanksby

The aim of this study was to examine the effect of endurance training on skeletal muscle phospholipid molecular species from high-fat fed rats. Twelve female Sprague- Dawley rats were fed a high-fat diet (78.1% energy). The rats were randomly divided into two groups, a sedentary control group and a trained group (125 min of treadmill running at 8 m/min, 4 days/wk for 4 weeks). Forty-eight hours after their last training bout phospholipids were extracted from the red and white vastus lateralis and analyzed by electrospray-ionization mass spectrometry. Exercise training was associated with significant alterations in the relative abundance of a …

Synergy Sted Multicolor 3d-Easysted Excel File, George Mcnamara

George McNamara

Excel 2007 file with calculations for multicolor 3D-EasySTED segmented waveplates (SWP) for fluorescence nanoscopy.

Additional explanation in my Introduction statement and in the Excel file. The Excel file uses an interpolation function available from XlXtrFun (http://www.xlxtrfun.com/XlXtrFun/XlXtrFun.htm). My thanks to Goetz Zinner, www.B-Halle.de, for communicating to me that a single 3D-EasySTED segmented waveplate can be used with multiple excitation and depletion wavelengths.

Please give Goetz Zinner full credit for recognizing that STED segmented waveplate can be manufactured to handle multiple excitation and depletion wavelengths. Different sets of wavelengths can be generated by choosing appropriate thickness quartz substrates for manufacturing the SWP. …