William wimley

Cyclic peptides that inhibit protein-protein interactions have significant advantages over linear peptides and small molecules for modulating cellular signaling networks in cancer and other diseases. However, the permeability barrier of the plasma membrane remains a formidable obstacle to the development of cyclic peptides into applicable drugs. Here, we test the ability of a family of synthetically evolved spontaneous membrane translocating peptides (SMTPs) to deliver phalloidin, a representative bioactive cyclic peptide, to the cytosol of human cells in culture...

Chimeric antigen receptor (CAR)-T cells have demonstrated clinical potential, but current receptors still need improvements to be successful against chronic HIV infection. In this study, we address some requirements of CAR motifs for strong surface expression of a novel anti-HIV CAR by evaluating important elements in the extracellular, hinge, and transmembrane (TM) domains. When combining a truncated CD4 extracellular domain and CD8α hinge/TM, the novel CAR did not express extracellularly but was detectable intracellularly...

We present a simple, spreadsheet-based method to determine the statistical significance of the difference between any two arbitrary curves. This modified Chi-squared method addresses two scenarios: A single measurement at each point with known standard deviation, or multiple measurements at each point averaged to produce a mean and standard error. The method includes an essential correction for the deviation from normality in measurements with small sample size, which are typical in biomedical sciences. Statistical significance is determined without regard to the functionality of the curves, or the signs of the differences...

Nanopore sensors have shown great utility in nucleic acid detection and sequencing approaches. Recent studies also indicate that current signatures produced by peptide-nanopore interactions can distinguish high purity peptide mixtures, but the utility of nanopore sensors in clinical applications still needs to be explored due to the inherent complexity of clinical specimens. To fill this gap between research and clinical nanopore applications, we describe a methodology to select peptide biomarkers suitable for use in an immunoprecipitation-coupled nanopore (IP-NP) assay, based on their pathogen specificity, antigenicity, charge, water solubility and ability to produce a characteristic nanopore interaction signature...

Here, we describe the continued synthetic molecular evolution of a lineage of host-compatible antimicrobial peptides (AMP) intended for the treatment of wounds infected with drug-resistant, biofilm-forming bacteria. The peptides tested are variants of an evolved AMP called d-amino acid CONsensus with Glycine Absent (d-CONGA), which has excellent antimicrobial activities in vitro and in vivo . In this newest generation of rational d-CONGA variants, we tested multiple sequence-structure-function hypotheses that had not been tested in previous generations...

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Membrane-active peptides play an essential role in many living organisms and their immune systems and counter many infectious diseases. Many have dual or multiple mechanisms and can synergize with other molecules, like peptides, proteins, and small molecules. Although membrane-active peptides have been intensively studied in the past decades and more than 3500 sequences have been identified, only a few received approvals from the U.S. Food and Drug Administration. In this review, we investigated all the peptide therapeutics that have entered the market or were subjected to preclinical and clinical studies to understand how they succeeded...

The receptor tyrosine kinase (RTK) EphA2 is expressed in epithelial and endothelial cells and controls the assembly of cell-cell junctions. EphA2 has also been implicated in many diseases, including cancer. Unlike most RTKs, which signal predominantly as dimers, EphA2 readily forms higher order oligomers upon ligand binding. Here, we investigated if a correlation exists between EphA2 signaling properties and the size of the EphA2 oligomers induced by multiple ligands, including the widely used ephrinA1-Fc ligand, the soluble monomeric m-ephrinA1, and novel engineered peptide ligands...

Gram-negative bacteria belonging to the genus Burkholderia are remarkably resistant to broad-spectrum, cationic, antimicrobial peptides (AMPs). It has been proposed that this innate resistance is related to changes in the outer membrane lipopolysaccharide (OM LPS), including the constitutive, essential modification of outer membrane Lipid A phosphate groups with cationic 4-amino-4-deoxy-arabinose. This modification reduces the overall negative charge on the OM LPS which may change the OM structure and reduce the binding, accumulation, and permeation of cationic AMPs...

During the 2013-2016 West African (WA) Ebola virus (EBOV) outbreak, severe gastrointestinal symptoms were common in patients and associated with poor outcome. Delta peptide is a conserved product of post-translational processing of the abundant EBOV soluble glycoprotein (sGP). The murine ligated ileal loop model was used to demonstrate that delta peptide is a potent enterotoxin. Dramatic intestinal fluid accumulation follows injection of biologically relevant amounts of delta peptide into ileal loops, along with gross alteration of villous architecture and loss of goblet cells...

Rational design and optimization of cell penetrating peptides (CPPs) is difficult to accomplish because of the lack of quantitative sequence-structure-function rules describing the activity and because of the complex, poorly understood mechanisms of CPPs. Synthetic molecular evolution is a powerful method to identify gain-of-function cell penetrating peptide variants in this situation. Synthetic molecular evolution requires the design and synthesis of iterative, knowledge-based peptide libraries and the screening of such libraries in complex orthogonal cell-based screens for improved activity...

BACKGROUND: Biofilms are microbial communities surrounded by a self-produced extracellular matrix which protects them from environmental stress. Bacteria within biofilms are 10- to 1000-fold more resistant to antibiotics, making it challenging but imperative to develop new therapeutics that can disperse biofilms and eradicate infection. Gram-negative bacteria produce outer membrane vesicles (OMV) that play critical roles in communication, genetic exchange, cargo delivery, and pathogenesis...

Growing evidence indicates that prorenin receptor (PRR) is upregulated in collecting duct (CD) of diabetic kidney. Prorenin is secreted by the principal CD cells, and is the natural ligand of the PRR. PRR activation stimulates fibrotic factors, including fibronectin, collagen, and transforming growth factor-β (TGF-β) contributing to tubular fibrosis. However, whether high glucose (HG) contributes to this effect is unknown. We tested the hypothesis that HG increases the abundance of PRR at the plasma membrane of the CD cells, thus contributing to the stimulation of downstream fibrotic factors, including TGF-β, collagen I, and fibronectin...

Peptides that form nanoscale pores in lipid bilayers have potential applications in triggered release, but only if their selectivity for target synthetic membranes over bystander biomembranes can be optimized. Previously, we identified a novel family of α-helical pore-forming peptides called "macrolittins", which release macromolecular cargoes from phosphatidylcholine (PC) liposomes at concentrations as low as 1 peptide per 1000 lipids. In this work, we show that macrolittins have no measurable cytolytic activity against multiple human cell types even at high peptide concentration...

Hepatitis C virus (HCV) infection promotes autophagic degradation of viral replicative intermediates for sustaining replication and spread. The excessive activation of autophagy can induce cell death and terminate infection without proper regulation. A prior publication from this laboratory showed that an adaptive cellular response to HCV microbial stress inhibits autophagy through beclin 1 degradation. The mechanisms of how secretory and degradative autophagy are regulated during persistent HCV infection is unknown...

The use of designed antimicrobial peptides as drugs has been impeded by the absence of simple sequence-structure-function relationships and design rules. The likely cause is that many of these peptides permeabilize membranes via highly disordered, heterogeneous mechanisms, forming aggregates without well-defined tertiary or secondary structure. We suggest that the combination of high-throughput library screening with atomistic computer simulations can successfully address this challenge by tuning a previously developed general pore-forming peptide into a selective pore-former for different lipid types...

Peptides that self-assemble into nanometer-sized pores in lipid bilayers could have utility in a variety of biotechnological and clinical applications if we can understand their physical chemical properties and learn to control their membrane selectivity. To empower such control, we have used synthetic molecular evolution to identify the pH-dependent delivery peptides, a family of peptides that assemble into macromolecule-sized pores in membranes at low peptide concentration but only at pH < ∼6. Further advancements will also require better selectivity for specific membranes...

Numerous peptides inhibit the entry of enveloped viruses into cells. Some of these peptides have been shown to inhibit multiple unrelated viruses. We have suggested that such broad-spectrum antiviral peptides share a property called interfacial activity; they are somewhat hydrophobic and amphipathic, with a propensity to interact with the interfacial zones of lipid bilayer membranes. In this study, we further tested the hypothesis that such interfacial activity is a correlate of broad-spectrum antiviral activity...

The synthetically evolved pH-dependent delivery (pHD) peptides are a unique family of peptides that bind to membranes, fold into α-helices, and form macromolecule-sized pores at low concentration in response to acidic pH < 6. They have potential applications in drug delivery and tumor targeting but their activity and membrane selectivity must be optimized. Here, we develop a better understanding of how pHD peptide activity in membranes can be modulated by increasing hydrophobicity and membrane binding without changing amino acid sequence...

The path to our modern understanding of the structure of the lipid bilayer membrane is a long one that can be traced from today perhaps as far back as Benjamin Franklin in the eighteenth century. Here, I provide a personal account of one of the important steps in that path, the description of the "Complete Structure" of a hydrated, fluid phase dioleoyl phosphatidylcholine bilayer by the joint refinement of neutron and X-ray diffraction data by Stephen White and his colleagues.