Preparation and characterization of the extracellular domain of human Sid-1
In C. elegans, the cell surface protein Sid-1 imports extracellular dsRNA into the cytosol of most non-neuronal cells, enabling systemic spread of RNA interference (RNAi) throughout the worm. Sid-1 homologs are found in many other animals, although for most a function for the protein has not yet bee...
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| creator | Pratt, Ashley J Rambo, Robert P Lau, Pick-Wei MacRae, Ian J |
| description | In C. elegans, the cell surface protein Sid-1 imports extracellular dsRNA into the cytosol of most non-neuronal cells, enabling systemic spread of RNA interference (RNAi) throughout the worm. Sid-1 homologs are found in many other animals, although for most a function for the protein has not yet been established. Sid-1 proteins are composed of an N-terminal extracellular domain (ECD) followed by 9-12 predicted transmembrane regions. We developed a baculovirus system to express and purify the ECD of the human Sid-1 protein SidT1. Recombinant SidT1 ECD is glycosylated and spontaneously assembles into a stable and discrete tetrameric structure. Electron microscopy (EM) and small angle x-ray scattering (SAXS) studies reveal that the SidT1 ECD tetramer is a compact, puck-shaped globular particle, which we hypothesize may control access of dsRNA to the transmembrane pore. These characterizations provide inroads towards understanding the mechanism of this unique RNA transport system from structural prospective. |
| doi_str_mv | 10.1371/journal.pone.0033607 |
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These characterizations provide inroads towards understanding the mechanism of this unique RNA transport system from structural prospective.</description><subject>Access control</subject><subject>Animals</subject><subject>Baculoviridae - genetics</subject><subject>Baculovirus</subject><subject>BASIC BIOLOGICAL SCIENCES</subject><subject>Biology</subject><subject>Caenorhabditis elegans</subject><subject>Cell surface</subject><subject>Crystallography</subject><subject>Cytosol</subject><subject>Data analysis</subject><subject>Double-stranded RNA</subject><subject>Electron microscopy</subject><subject>Extracellular Space - metabolism</subject><subject>Gene expression</subject><subject>Genes</subject><subject>Glycosylation</subject><subject>Homology</subject><subject>Humans</subject><subject>Imports</subject><subject>Insects</subject><subject>Membrane Transport Proteins - chemistry</subject><subject>Membrane Transport Proteins - genetics</subject><subject>Membrane Transport Proteins - isolation & purification</subject><subject>Membrane Transport Proteins - 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Sid-1 homologs are found in many other animals, although for most a function for the protein has not yet been established. Sid-1 proteins are composed of an N-terminal extracellular domain (ECD) followed by 9-12 predicted transmembrane regions. We developed a baculovirus system to express and purify the ECD of the human Sid-1 protein SidT1. Recombinant SidT1 ECD is glycosylated and spontaneously assembles into a stable and discrete tetrameric structure. Electron microscopy (EM) and small angle x-ray scattering (SAXS) studies reveal that the SidT1 ECD tetramer is a compact, puck-shaped globular particle, which we hypothesize may control access of dsRNA to the transmembrane pore. These characterizations provide inroads towards understanding the mechanism of this unique RNA transport system from structural prospective.</abstract><cop>United States</cop><pub>Public Library of Science</pub><pmid>22509261</pmid><doi>10.1371/journal.pone.0033607</doi><tpages>e33607</tpages><oa>free_for_read</oa></addata></record> |
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| subjects | Access control Animals Baculoviridae - genetics Baculovirus BASIC BIOLOGICAL SCIENCES Biology Caenorhabditis elegans Cell surface Crystallography Cytosol Data analysis Double-stranded RNA Electron microscopy Extracellular Space - metabolism Gene expression Genes Glycosylation Homology Humans Imports Insects Membrane Transport Proteins - chemistry Membrane Transport Proteins - genetics Membrane Transport Proteins - isolation & purification Membrane Transport Proteins - metabolism Models, Molecular Molecular biology Nematodes Physiology Potassium Protein Folding Protein Multimerization Protein Processing, Post-Translational protein structure protein structure prediction Protein Structure, Quaternary Protein Structure, Tertiary Proteins Pyrococcus horikoshii recombinant protein Ribonucleic acid RNA RNA structure RNA transport RNA-mediated interference Sequence Homology, Amino Acid small angle scattering Small angle X ray scattering Solubility Spodoptera - cytology Trends Vertebrates Viral infections X ray scattering |
| title | Preparation and characterization of the extracellular domain of human Sid-1 |
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