Controlling the Size and Shape of Polypeptide Colloidal Particles: Temperature Dependence of Particle Formation

A promising approach for developing new drug delivery vehicles is by using stimuli responsive hydrogel nanoparticles. Polypeptide surfactants designed in our lab have been shown to form micellar particles of varying sizes and shapes depending on the solution salt concentration. These responsive polypeptide surfactants consist of a small charged protein domain (foldon) with three elastin-like polypeptide (ELP) chains forming a three-armed star polymer. The size and shape of the micelles they form is dependent on the ratio of total ELP volume to head group area. By introducing linear ELP into the ELP-foldon solution, the total volume of ELP in the aggregate would be increased if the linear ELP is incorporated in the micelle. This method could control the particle size and shape. To determine if the linear and three-armed ELPs co-assemble, we have observed aggregation as a function of temperature using turbidity measurements in a UV-vis spectrometer. We have found that higher concentrations of linear ELP increases the difference in transition temperature between the linear and three-armed ELP. At these higher ratios, the linear ELP aggregates prior to micelle formation. When the ELP-foldon subsequently passes through its critical micelle temperature, they break down the linear ELP aggregates resulting in smaller colloidal emulsions. Light scattering will be used to characterize the size and shape of these aggregates.https://engagedscholarship.csuohio.edu/u_poster_2013/1016/thumbnail.jp

Controlling the Size and Shape of Polypeptide Colloidal Particles: Temperature Dependence of Particle Formation

A promising approach for developing new drug delivery vehicles is by using stimuli responsive hydrogel nanoparticles. Polypeptide surfactants designed in our lab have been shown to form micellar particles of varying sizes and shapes depending on the solution salt concentration. These responsive polypeptide surfactants consist of a small charged protein domain (foldon) with three elastin-like polypeptide (ELP) chains forming a three-armed star polymer. The size and shape of the micelles they form is dependent on the ratio of total ELP volume to head group area. By introducing linear ELP into the ELP-foldon solution, the total volume of ELP in the aggregate would be increased if the linear ELP is incorporated in the micelle. This method could control the particle size and shape. To determine if the linear and three-armed ELPs co-assemble, we have observed aggregation as a function of temperature using turbidity measurements in a UV-vis spectrometer. We have found that higher concentrations of linear ELP increases the difference in transition temperature between the linear and three-armed ELP. At these higher ratios, the linear ELP aggregates prior to micelle formation. When the ELP-foldon subsequently passes through its critical micelle temperature, they break down the linear ELP aggregates resulting in smaller colloidal emulsions. Light scattering will be used to characterize the size and shape of these aggregates.https://engagedscholarship.csuohio.edu/u_poster_2013/1016/thumbnail.jp

Periodic Operation of a Dynamic DNA Origami Structure Utilizing the Hydrophilic–Hydrophobic Phase‐Transition of Stimulus‐Sensitive Polypeptides

Dynamic DNA nanodevices are designed to perform structure‐encoded motion actuated by a variety of different physicochemical stimuli. In this context, hybrid devices utilizing other components than DNA have the potential to considerably expand the library of functionalities. Here, the reversible reconfiguration of a DNA origami structure using the stimulus sensitivity of elastin‐like polypeptides is reported. To this end, a rectangular sheet made using the DNA origami technique is functionalized with these peptides and by applying changes in salt concentration the hydrophilic–hydrophobic phase transition of these peptides actuate the folding of the structure. The on‐demand and reversible switching of the rectangle is driven by externally imposed temperature oscillations and appears at specific transition temperatures. Using transmission electron microscopy, it is shown that the structure exhibits distinct conformational states with different occupation probabilities, which are dependent on structure‐intrinsic parameters such as the local number and the arrangement of the peptides on the rectangle. It is also shown through ensemble fluorescence resonance energy transfer spectroscopy that the transition temperature and thus the thermodynamics of the rectangle‐peptide system depends on the stimuli salt concentration and temperature, as well as on the intrinsic parameters

Integrated modelling of cost-effective siting and operation of flow-control infrastructure for river ecosystem conservation

Wetland and floodplain ecosystems along many regulated rivers are highly stressed, primarily due to a lack of environmental flows of appropriate magnitude, frequency, duration, and timing to support ecological functions. In the absence of increased environmental flows, the ecological health of river ecosystems can be enhanced by the operation of existing and new flow-control infrastructure (weirs and regulators) to return more natural environmental flow regimes to specific areas. However, determining the optimal investment and operation strategies over time is a complex task due to several factors including the multiple environmental values attached to wetlands, spatial and temporal heterogeneity and dependencies, nonlinearity, and time-dependent decisions. This makes for a very large number of decision variables over a long planning horizon. The focus of this paper is the development of a nonlinear integer programming model that accommodates these complexities. The mathematical objective aims to return the natural flow regime of key components of river ecosystems in terms of flood timing, flood duration, and interflood period. We applied a 2-stage recursive heuristic using tabu search to solve the model and tested it on the entire South Australian River Murray floodplain. We conclude that modern meta-heuristics can be used to solve the very complex nonlinear problems with spatial and temporal dependencies typical of environmental flow allocation in regulated river ecosystems. The model has been used to inform the investment in, and operation of, flow-control infrastructure in the South Australian River Murray.<br /

Swift and Fermi observations of the early afterglow of the short Gamma-Ray Burst 090510

We present the observations of GRB090510 performed by the Fermi Gamma-Ray Space Telescope and the Swift observatory. This is a bright, short burst that shows an extended emission detected in the GeV range. Furthermore, its optical emission initially rises, a feature so far observed only in long bursts, while the X-ray flux shows an initial shallow decrease, followed by a steeper decay. This exceptional behavior enables us to investigate the physical properties of the GRB outflow, poorly known in short bursts. We discuss internal shock and external shock models for the broadband energy emission of this object.Comment: Comments: Submitted to ApJ Letters. Contact Authors: Massimiliano De Pasquale ([email protected]), Mathew Page ([email protected]), Kenji Toma ([email protected]), Veronique Pelassa ([email protected]). Minor change in the authorlis

The Transcription Factor AmrZ Utilizes Multiple DNA Binding Modes to Recognize Activator and Repressor Sequences of Pseudomonas aeruginosa Virulence Genes

AmrZ, a member of the Ribbon-Helix-Helix family of DNA binding proteins, functions as both a transcriptional activator and repressor of multiple genes encoding Pseudomonas aeruginosa virulence factors. The expression of these virulence factors leads to chronic and sustained infections associated with worsening prognosis. In this study, we present the X-ray crystal structure of AmrZ in complex with DNA containing the repressor site, amrZ1. Binding of AmrZ to this site leads to auto-repression. AmrZ binds this DNA sequence as a dimer-of-dimers, and makes specific base contacts to two half sites, separated by a five base pair linker region. Analysis of the linker region shows a narrowing of the minor groove, causing significant distortions. AmrZ binding assays utilizing sequences containing variations in this linker region reveals that secondary structure of the DNA, conferred by the sequence of this region, is an important determinant in binding affinity. The results from these experiments allow for the creation of a model where both intrinsic structure of the DNA and specific nucleotide recognition are absolutely necessary for binding of the protein. We also examined AmrZ binding to the algD promoter, which results in activation of the alginate exopolysaccharide biosynthetic operon, and found the protein utilizes different interactions with this site. Finally, we tested the in vivo effects of this differential binding by switching the AmrZ binding site at algD, where it acts as an activator, for a repressor binding sequence and show that differences in binding alone do not affect transcriptional regulation

Computational Prediction and Experimental Verification of New MAP Kinase Docking Sites and Substrates Including Gli Transcription Factors

In order to fully understand protein kinase networks, new methods are needed to identify regulators and substrates of kinases, especially for weakly expressed proteins. Here we have developed a hybrid computational search algorithm that combines machine learning and expert knowledge to identify kinase docking sites, and used this algorithm to search the human genome for novel MAP kinase substrates and regulators focused on the JNK family of MAP kinases. Predictions were tested by peptide array followed by rigorous biochemical verification with in vitro binding and kinase assays on wild-type and mutant proteins. Using this procedure, we found new ‘D-site’ class docking sites in previously known JNK substrates (hnRNP-K, PPM1J/PP2Czeta), as well as new JNK-interacting proteins (MLL4, NEIL1). Finally, we identified new D-site-dependent MAPK substrates, including the hedgehog-regulated transcription factors Gli1 and Gli3, suggesting that a direct connection between MAP kinase and hedgehog signaling may occur at the level of these key regulators. These results demonstrate that a genome-wide search for MAP kinase docking sites can be used to find new docking sites and substrates