Modular assembly of a protein nanotriangle using orthogonally interacting coiled coils

Synthetic protein assemblies that adopt programmed shapes would support many applications in nanotechnology. We used a rational design approach that exploits the modularity of orthogonally interacting coiled coils to create a self-assembled protein nanotriangle. Coiled coils have frequently been used to construct nanoassemblies and materials, but rarely with successful prior specification of the resulting structure. We designed a heterotrimer from three pairs of heterodimeric coiled coils that mediate specific interactions while avoiding undesired crosstalk. Non-associating pairs of coiled-coil units were strategically fused to generate three chains that were predicted to preferentially form the heterotrimer, and a rational annealing proc ess led to the desired oligomer. Extensive biophysical characterization and modeling support the formation of a molecular triangle, which is a shape distinct from naturally occurring supramolecular nanostructures. Our approach can be extended to design more complex nanostructures using additional coiled-coil modules, other protein parts, or templated surfaces

Taking aim at Mer and Axl receptor tyrosine kinases as novel therapeutic targets in solid tumors

Axl and/or Mer expression correlates with poor prognosis in several cancers. Until recently, the specific role of these receptor tyrosine kinases (RTKs) in the development and progression of cancer remained unexplained. Studies demonstrating that Axl and Mer contribute to mechanisms of cell survival, migration, invasion, metastasis, and chemosensitivity justify further investigation of Axl and Mer as novel therapeutic targets in cancer

The MCL-1 BH3 helix is an exclusive MCL-1 inhibitor and apoptosis sensitizer

available in PMC 2011 February 3.MCL-1 has emerged as a major oncogenic and chemoresistance factor. A screen of stapled peptide helices identified the MCL-1 BH3 domain as selectively inhibiting MCL-1 among the related anti-apoptotic Bcl-2 family members, providing insights into the molecular determinants of binding specificity and a new approach for sensitizing cancer cells to apoptosis.National Institutes of Health (U.S.) (NIH award 5RO1GM084181)National Institutes of Health (U.S.) (NIH grant 5P01CA92625)National Institutes of Health (U.S.) (Ruth L. Kirschstein National Research Service Award 1F31CA144566)Burroughs Wellcome Fund (Career Award

Changes in neuronal activation patterns in response to androgen deprivation therapy: a pilot study

<p>Abstract</p> <p>Background</p> <p>A common treatment option for men with prostate cancer is androgen deprivation therapy (ADT). However, men undergoing ADT may experience physical side effects, changes in quality of life and sometimes psychiatric and cognitive side effects.</p> <p>Methods</p> <p>In this study, hormone naïve patients without evidence of metastases with a rising PSA were treated with nine months of ADT. Functional magnetic resonance imaging (fMRI) of the brain during three visuospatial tasks was performed at baseline prior to treatment and after nine months of ADT in five subjects. Seven healthy control patients, underwent neuroimaging at the same time intervals.</p> <p>Results</p> <p>ADT patients showed reduced, task-related BOLD-fMRI activation during treatment that was not observed in control subjects. Reduction in activation in right parietal-occipital regions from baseline was observed during recall of the spatial location of objects and mental rotation.</p> <p>Conclusions</p> <p>Findings, while preliminary, suggest that ADT reduces task-related neural activation in brain regions that are involved in mental rotation and accurate recall of spatial information.</p

The impact of donor and recipient common clinical and genetic variation on estimated glomerular filtration rate in a European renal transplant population

Genetic variation across the HLA is known to influence renal‐transplant outcome. However, the impact of genetic variation beyond the HLA is less clear. We tested the association of common genetic variation and clinical characteristics, from both the donor and recipient, with post‐transplant eGFR at different time‐points, out to 5‐years post‐transplantation. We conducted GWAS meta‐analyses across 10,844 donors and recipients from five European ancestry cohorts. We also analysed the impact of polygenic risk scores (PRS), calculated using genetic variants associated with non‐transplant eGFR, on post‐transplant eGFR. PRS calculated using the recipient genotype alone, as well as combined donor and recipient genotypes were significantly associated with eGFR at 1‐year post‐transplant. 32% of the variability in eGFR at 1‐year post‐transplant was explained by our model containing clinical covariates (including weights for death/graft‐failure), principal components and combined donor‐recipient PRS, with 0.3% contributed by the PRS. No individual genetic variant was significantly associated with eGFR post‐transplant in the GWAS. This is the first study to examine PRS, composed of variants that impact kidney function in the general population, in a post‐transplant context. Despite PRS being a significant predictor of eGFR post‐transplant, the effect size of common genetic factors is limited compared to clinical variables

Enteric Pathogens in Stored Drinking Water and on Caregiver's Hands in Tanzanian Households with and without Reported Cases of Child Diarrhea.

Diarrhea is one of the leading causes of mortality in young children. Diarrheal pathogens are transmitted via the fecal-oral route, and for children the majority of this transmission is thought to occur within the home. However, very few studies have documented enteric pathogens within households of low-income countries. The presence of molecular markers for three enteric viruses (enterovirus, adenovirus, and rotavirus), seven Escherichia coli virulence genes (ECVG), and human-specific Bacteroidales was assessed in hand rinses and household stored drinking water in Bagamoyo, Tanzania. Using a matched case-control study design, we examined the relationship between contamination of hands and water with these markers and child diarrhea. We found that the presence of ECVG in household stored water was associated with a significant decrease in the odds of a child within the home having diarrhea (OR = 0.51; 95% confidence interval 0.27-0.93). We also evaluated water management and hygiene behaviors. Recent hand contact with water or food was positively associated with detection of enteric pathogen markers on hands, as was relatively lower volumes of water reportedly used for daily hand washing. Enteropathogen markers in stored drinking water were more likely found among households in which the markers were also detected on hands, as well as in households with unimproved water supply and sanitation infrastructure. The prevalence of enteric pathogen genes and the human-specific Bacteroidales fecal marker in stored water and on hands suggests extensive environmental contamination within homes both with and without reported child diarrhea. Better stored water quality among households with diarrhea indicates caregivers with sick children may be more likely to ensure safe drinking water in the home. Interventions to increase the quantity of water available for hand washing, and to improve food hygiene, may reduce exposure to enteric pathogens in the domestic environment

Peptide Ligands for Pro-survival Protein Bfl-1 from Computationally Guided Library Screening

Pro-survival members of the Bcl-2 protein family inhibit cell death by binding short helical BH3 motifs in pro-apoptotic proteins. Mammalian pro-survival proteins Bcl-x[subscript L], Bcl-2, Bcl-w, Mcl-1, and Bfl-1 bind with varying affinities and specificities to native BH3 motifs, engineered peptides, and small molecules. Biophysical studies have determined interaction patterns for these proteins, particularly for the most-studied family members Bcl-x[subscript L] and Mcl-1. Bfl-1 is a pro-survival protein implicated in preventing apoptosis in leukemia, lymphoma, and melanoma. Although Bfl-1 is a promising therapeutic target, relatively little is known about its binding preferences. We explored the binding of Bfl-1 to BH3-like peptides by screening a peptide library that was designed to sample a high degree of relevant sequence diversity. Screening using yeast-surface display led to several novel high-affinity Bfl-1 binders and to thousands of putative binders identified through deep sequencing. Further screening for specificity led to identification of a peptide that bound to Bfl-1 with K[subscript d] < 1 nM and very slow dissociation from Bfl-1 compared to other pro-survival Bcl-2 family members. A point mutation in this sequence gave a peptide with ~50 nM affinity for Bfl-1 that was selective for Bfl-1 in equilibrium binding assays. Analysis of engineered Bfl-1 binders deepens our understanding of how the binding profiles of pro-survival proteins differ and may guide the development of targeted Bfl-1 inhibitors.National Institute of General Medical Sciences (U.S.) (Award GM084181)National Institute of General Medical Sciences (U.S.) (Award P50-GM68762

Identification of a Novel Class of Farnesylation Targets by Structure-Based Modeling of Binding Specificity

Farnesylation is an important post-translational modification catalyzed by farnesyltransferase (FTase). Until recently it was believed that a C-terminal CaaX motif is required for farnesylation, but recent experiments have revealed larger substrate diversity. In this study, we propose a general structural modeling scheme to account for peptide binding specificity and recapitulate the experimentally derived selectivity profile of FTase in vitro. In addition to highly accurate recovery of known FTase targets, we also identify a range of novel potential targets in the human genome, including a new substrate class with an acidic C-terminal residue (CxxD/E). In vitro experiments verified farnesylation of 26/29 tested peptides, including both novel human targets, as well as peptides predicted to tightly bind FTase. This study extends the putative range of biological farnesylation substrates. Moreover, it suggests that the ability of a peptide to bind FTase is a main determinant for the farnesylation reaction. Finally, simple adaptation of our approach can contribute to more accurate and complete elucidation of peptide-mediated interactions and modifications in the cell

Association of genetic variation with systolic and diastolic blood pressure among African Americans: the Candidate Gene Association Resource study

The prevalence of hypertension in African Americans (AAs) is higher than in other US groups; yet, few have performed genome-wide association studies (GWASs) in AA. Among people of European descent, GWASs have identified genetic variants at 13 loci that are associated with blood pressure. It is unknown if these variants confer susceptibility in people of African ancestry. Here, we examined genome-wide and candidate gene associations with systolic blood pressure (SBP) and diastolic blood pressure (DBP) using the Candidate Gene Association Resource (CARe) consortium consisting of 8591 AAs. Genotypes included genome-wide single-nucleotide polymorphism (SNP) data utilizing the Affymetrix 6.0 array with imputation to 2.5 million HapMap SNPs and candidate gene SNP data utilizing a 50K cardiovascular gene-centric array (ITMAT-Broad-CARe [IBC] array). For Affymetrix data, the strongest signal for DBP was rs10474346 (P= 3.6 × 10−8) located near GPR98 and ARRDC3. For SBP, the strongest signal was rs2258119 in C21orf91 (P= 4.7 × 10−8). The top IBC association for SBP was rs2012318 (P= 6.4 × 10−6) near SLC25A42 and for DBP was rs2523586 (P= 1.3 × 10−6) near HLA-B. None of the top variants replicated in additional AA (n = 11 882) or European-American (n = 69 899) cohorts. We replicated previously reported European-American blood pressure SNPs in our AA samples (SH2B3, P= 0.009; TBX3-TBX5, P= 0.03; and CSK-ULK3, P= 0.0004). These genetic loci represent the best evidence of genetic influences on SBP and DBP in AAs to date. More broadly, this work supports that notion that blood pressure among AAs is a trait with genetic underpinnings but also with significant complexit