Size-dependent control of the binding of biotinylated proteins to streptavidin using a polymer shield

Many medical and biotechnological processes rely on controlling and manipulating the molecular-recognition capabilities of proteins. This can be achieved using small molecules capable of competing for protein binding or by changing environmental parameters that affect protein structure and hence binding. An alternative is provided by stimuli-responsive polymers that change reversibly from a water-soluble expanded coil to a water-insoluble collapsed globule upon small changes in temperature, pH or light intensity: when attached to proteins in the vicinity of their binding sites, they reversibly block and release small ligands. Here we show how this approach can be extended to achieve size-selective binding of large, macromolecular ligands. We use the thermally responsive polymer poly(N,N-diethylacrylamide) (PDEAAm), and attach it to the protein streptavidin approximately 20 A from the binding site for biotinylated proteins. Below the lower critical solution temperature of PDEAAm, the polymer is in its extended state and acts as a 'shield' to block the binding of large biotinylated proteins; above this temperature, it collapses and exposes the binding site, thereby allowing binding. We find that the degree of shielding depends on both the size of the biotinylated protein and the size of PDEAAm, suggesting that 'smart' polymer shields could be tailored to achieve a wide range of size-dependent ligand discrimination for use in affinity separations, biosensors and diagnostics technologies.     

Pregnancy following segmental isthmic reversal of the rabbit oviduct

Summary Microsurgical reversal of a segment of rabbit proximal tubal isthmus has been followed by normal pregnancy in the first two animals to undergo the procedure. Establishment of pregnancy despite radical modification of the oviduct furnishes the opportunity to gain new insights into the mechanisms controlling tubal ovum transport and emphasizes the evolving feasibility and importance of tuboplastic microsurgery both as a research tool and clinical procedure.This study was partly supported by an N.I.H. Institutional Research Grant.Supported by the Rockefeller Foundation.     

Design of Smart Polymer-Protein Conjugates and Smart Magnetic Nanoparticles for Use in Microfluidic Diagnostic Assays

1 Results In this talk,I will describe the design,synthesis and application of smart polymers for use in microfluidic diagnostic devices.We are synthesizing a variety of temperature- and pH-responsive polymers using RAFT living free radical polymerization techniques.This allows us to control molecular weight and to achieve a narrow MW distribution of the polymers. Furthermore,RAFT polymers have reactive end groups that are used to conjugate the polymers to proteins.We are also using those groups to bind...     

Dynamic molecular processes mediate cellular mechanotransduction

Cellular responses to mechanical forces are crucial in embryonic development and adult physiology, and are involved in numerous diseases, including atherosclerosis, hypertension, osteoporosis, muscular dystrophy, myopathies and cancer. These responses are mediated by load-bearing subcellular structures, such as the plasma membrane, cell-adhesion complexes and the cytoskeleton. Recent work has demonstrated that these structures are dynamic, undergoing assembly, disassembly and movement, even when ostensibly stable. An emerging insight is that transduction of forces into biochemical signals occurs within the context of these processes. This framework helps to explain how forces of varying strengths or dynamic characteristics regulate distinct signalling pathways.     

Molecular cloning establishes proenkephalin as precursor of enkephalin-containing peptides

Molecular cloning and DNA sequencing have yielded considerable structural information about proenkephalin. All previously characterized intermediate peptides of the enkephalin pathways in bovine adrenal medulla have now been aligned into an unambiguous primary structure. Two basic amino acid residues serve as processing signals for release of each of the different components.