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Title: LDRD final report on imaging self-organization of proteins in membranes by photocatalytic nano-tagging.

Abstract

We have developed a new nanotagging technology for detecting and imaging the self-organization of proteins and other components of membranes at nanometer resolution for the purpose of investigating cell signaling and other membrane-mediated biological processes. We used protein-, lipid-, or drug-bound porphyrin photocatalysts to grow in-situ nanometer-sized metal particles, which reveal the location of the porphyrin-labeled molecules by electron microscopy. We initially used photocatalytic nanotagging to image assembled multi-component proteins and to monitor the distribution of lipids and porphyrin labels in liposomes. For example, by exchanging the heme molecules in hemoproteins with a photocatalytic tin porphyrin, a nanoparticle was grown at each heme site of the protein. The result obtained from electron microscopy for a tagged multi-subunit protein such as hemoglobin is a symmetric constellation of a specific number of nanoparticle tags, four in the case of the hemoglobin tetramer. Methods for covalently linking photocatalytic porphyrin labels to lipids and proteins were also developed to detect and image the self-organization of lipids, protein-protein supercomplexes, and membrane-protein complexes. Procedures for making photocatalytic porphyrin-drug, porphyrin-lipid, and porphyrin-protein hybrids for non-porphyrin-binding proteins and membrane components were pursued and the first porphyrin-labeled lipids was investigated in liposomal membrane models. Our photocatalytic nanotagging technique maymore » ultimately allow membrane self-organization and cell signaling processes to be imaged in living cells. Fluorescence and plasmonic spectra of the tagged proteins might also provide additional information about protein association and membrane organization. In addition, a porphyrin-aspirin or other NSAID hybrid may be used to grow metal nanotags for the pharmacologically important COX enzymes in membranes so that the distribution of the protein can be imaged at the nanometer scale.« less

Authors:
; ; ; ;
Publication Date:
Research Org.:
Sandia National Laboratories
Sponsoring Org.:
USDOE
OSTI Identifier:
875973
Report Number(s):
SAND2005-6948
TRN: US200604%%280
DOE Contract Number:
AC04-94AL85000
Resource Type:
Technical Report
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; DISTRIBUTION; ELECTRON MICROSCOPY; ENZYMES; FLUORESCENCE; HEME; HEMOGLOBIN; LIPIDS; LIPOSOMES; MEMBRANES; MONITORS; PORPHYRINS; PROTEINS; RESOLUTION; SPECTRA; Cell membranes.; Protein-analysis.; Membranes.

Citation Formats

Zavadil, Kevin Robert, Shelnutt, John Allen, Sasaki, Darryl Yoshio, Song, Yujiang, and Medforth, Craig J. LDRD final report on imaging self-organization of proteins in membranes by photocatalytic nano-tagging.. United States: N. p., 2005. Web. doi:10.2172/875973.
Zavadil, Kevin Robert, Shelnutt, John Allen, Sasaki, Darryl Yoshio, Song, Yujiang, & Medforth, Craig J. LDRD final report on imaging self-organization of proteins in membranes by photocatalytic nano-tagging.. United States. doi:10.2172/875973.
Zavadil, Kevin Robert, Shelnutt, John Allen, Sasaki, Darryl Yoshio, Song, Yujiang, and Medforth, Craig J. Tue . "LDRD final report on imaging self-organization of proteins in membranes by photocatalytic nano-tagging.". United States. doi:10.2172/875973. https://www.osti.gov/servlets/purl/875973.
@article{osti_875973,
title = {LDRD final report on imaging self-organization of proteins in membranes by photocatalytic nano-tagging.},
author = {Zavadil, Kevin Robert and Shelnutt, John Allen and Sasaki, Darryl Yoshio and Song, Yujiang and Medforth, Craig J.},
abstractNote = {We have developed a new nanotagging technology for detecting and imaging the self-organization of proteins and other components of membranes at nanometer resolution for the purpose of investigating cell signaling and other membrane-mediated biological processes. We used protein-, lipid-, or drug-bound porphyrin photocatalysts to grow in-situ nanometer-sized metal particles, which reveal the location of the porphyrin-labeled molecules by electron microscopy. We initially used photocatalytic nanotagging to image assembled multi-component proteins and to monitor the distribution of lipids and porphyrin labels in liposomes. For example, by exchanging the heme molecules in hemoproteins with a photocatalytic tin porphyrin, a nanoparticle was grown at each heme site of the protein. The result obtained from electron microscopy for a tagged multi-subunit protein such as hemoglobin is a symmetric constellation of a specific number of nanoparticle tags, four in the case of the hemoglobin tetramer. Methods for covalently linking photocatalytic porphyrin labels to lipids and proteins were also developed to detect and image the self-organization of lipids, protein-protein supercomplexes, and membrane-protein complexes. Procedures for making photocatalytic porphyrin-drug, porphyrin-lipid, and porphyrin-protein hybrids for non-porphyrin-binding proteins and membrane components were pursued and the first porphyrin-labeled lipids was investigated in liposomal membrane models. Our photocatalytic nanotagging technique may ultimately allow membrane self-organization and cell signaling processes to be imaged in living cells. Fluorescence and plasmonic spectra of the tagged proteins might also provide additional information about protein association and membrane organization. In addition, a porphyrin-aspirin or other NSAID hybrid may be used to grow metal nanotags for the pharmacologically important COX enzymes in membranes so that the distribution of the protein can be imaged at the nanometer scale.},
doi = {10.2172/875973},
journal = {},
number = ,
volume = ,
place = {United States},
year = {Tue Nov 01 00:00:00 EST 2005},
month = {Tue Nov 01 00:00:00 EST 2005}
}

Technical Report:

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  • A new approach to the fabrication of porous, amorphous inorganic membranes using organic pore templates was investigated. The pore templates were a new family of hybrid organic-inorganic monomers. As background for membrane work, the monomers were polymerized by sol-gel techniques to make crosslinked polymers. Molecular modeling was used to create computer simulations of the materials and provide insight into their composites, were then converted into porous silicas using low temperature oxygen plasma techniques. A select few of the monomers were copolymerized with silica monomers to form non-porous thin films on mesoporous substrates. The films were converted into porous silica thinmore » films with thermal oxidations and the resulting membranes were tested for gas selectivities and flux.« less
  • We are currently exploring and developing a new statistical mechanics approach to designing self organizing and self assembling systems that is unique to SNL. The primary application target for this ongoing research is the development of new kinds of nanoscale components and hardware systems. However, a surprising out of the box connection to software development is emerging from this effort. With some amount of modification, the collective behavior physics ideas for enabling simple hardware components to self organize may also provide design methods for a new class of software modules. Large numbers of these relatively small software components, if designedmore » correctly, would be able to self assemble into a variety of much larger and more complex software systems. This self organization process would be steered to yield desired sets of system properties. If successful, this would provide a radical (disruptive technology) path to developing complex, high reliability software unlike any known today. The special work needed to evaluate this high risk, high payoff opportunity does not fit well into existing SNL funding categories, as it is well outside of the mainstreams of both conventional software development practices and the nanoscience research area that spawned it. We proposed a small LDRD effort aimed at appropriately generalizing these collective behavior physics concepts and testing their feasibility for achieving the self organization of large software systems. Our favorable results motivate an expanded effort to fully develop self-organizing software as a new technology.« less
  • This LDRD (Laboratory Directed Research and Development) project was funded for two years beginning in October 1992 (FY93) and was designed as a multidisciplinary approach to determining the structural and physical properties of C{sub 60} intercalated with various gases. The purpose of the study was to evaluate the relative permeation and diffusion of various gases with an ultimate goal of finding an effective filter for gas separations. A variety of probes including NMR, X-ray and neutron diffraction; IR spectroscopy, thermogravimetric analysis and mass spectroscopy were employed on C{sub 60} impregnated with a number of gases including O{sub 2}, N{sub 2},more » Ar, Ne, H{sub 2}, NO and CH{sub 4}. In order to increase the absorption and decrease the effective time constraints for bulk samples, these gases were intercalated into the C{sub 60} using pressures to several kbar. The results of these measurements which were quite encouraging for separation of O{sub 2} and N{sub 2} and for H{sub 2} from N{sub 2} led to 17 manuscripts which have been published in peer reviewed journals. The abstracts of these manuscripts are shown below along with a complete citation to the full text.« less
  • Using a combination of modeling and experimental work we have developed a new method for purifying water that uses less energy than conventional methods and that can be made selective for removing targeted contaminants. The method uses nanoporous membranes that are permselective for anion or cation transfer. Ion selectivity results from double layer overlap inside the pores such that they dominantly contain ions opposite in charge to the surface charge of the membrane. Membrane charge can be adjusted through functionalization. Experiments confirm membrane permselectivity and overall energy use less than that for conventional electrodialysis. The nanoporous membranes are used inmore » a conventional electrodialysis configuration and can be incorporated in existing electrodialysis systems without modification. The technology merits further development and testing in real systems, and could result in a significant reduction in water treatment costs.« less
  • The influence of ionic strength on the electrostatic interaction of viruses with environmentally relevant surfaces was determined for three viruses, MS2, Q{beta} and Norwalk. The environmental surface is modeled as charged Gouy-Chapman plane with and without a finite atomistic region (patch) of opposite charge. The virus is modeled as a particle comprised of ionizable amino acid residues in a shell surrounding a spherical RNA core of negative charge, these charges being compensated for by a Coulomb screening due to intercalated ions. Surface potential calculations for each of the viruses show excellent agreement with electrophoretic mobility and zeta potential measurements asmore » a function of pH. The results indicate that the electrostatic interaction between the virus and the planar surface, mitigated by the ionic strength of the solute, is dependent upon the spatial distribution of the amino acid residues in the different viruses. Specifically, the order of interaction energies with the patch (MS2 greatest at 5 mM; Norwalk greatest at 20 mM) is dependent upon the ionic strength of the fluid as a direct result of the viral coat amino acid distributions. We have developed an atomistic-scale method of calculation of the binding energy of viruses to surfaces including electrostatic, van der Waals, electron-overlap repulsion, surface charge polarization (images), and hydrophobic effects. The surface is treated as a Gouy-Chapman plane allowing inclusion of pH and ionic strength effects on the electrostatic potential at each amino acid charge. Van der Waals parameters are obtained from the DREIDING force field and from Hamaker constant measurements. We applied this method to the calculation of the Cowpea Mosaic Virus (CPMV), a negatively charged virus at a pH of 7.0, and find that the viral-gold surface interaction is very long range for both signs of surface potential, a result due to the electrostatic forces. For a negative (Au) surface potential of -0.05 volts, a nearly 4 eV barrier must be overcome to reach 1 nm from the surface.« less