Topological and Functional Analyses of the Triple-Gene-Block Protein 2 of Bamboo Mosaic Virus
| Autor: | Hsiu-Ting Hsu, 許琇婷 |
|---|---|
| Rok vydání: | 2009 |
| Druh dokumentu: | 學位論文 ; thesis |
| Popis: | 97 The triple gene block protein 2 (TGBp2) of Bamboo mosaic virus (BaMV) has been proposed to be a transmembrane protein; however, its features remain unclear. Here, we used biochemical approaches to determine its topological and biochemical properties. Our data reveal that TGBp2 is mainly associated with the endoplasmic reticulum membrane. The resistance of TGBp2 in proteoliposomes, prepared from both the BaMV-infected tissues and in vitro reconstitution system, to both chemical extraction and trypsin digestion confirmed that it is indeed an integral membrane protein. On the basis of the minor change in the size of the major stable TGBp2-derived tryptic fragment from the monomeric TGBp2, as well as the sensitivity of the cysteine residues at the C-terminal tail of TGBp2 to maleimide modification, we suggest that TGBp2 adopts a topology with both its short N- and C-terminal tails exposed to the outer surface of the endoplasmic reticulum. Moreover, TGBp2 is able to self-assemble as revealed by the ability to detect multimeric TGBp2 in the presence of crosslinker or oxidation agent. TGBp2 is thought to assist the movement of the viral ribonucleoprotein (RNP) complex by protein-protein or protein-RNA interactions. Using tyrosine fluorescence spectroscopy and UV-crosslinking assays, the TGBp2 solubilized with Triton X-100 was found to interact with viral RNA in a non-specific manner. These results raise the possibility that TGBp2 facilitates the intracellular delivery of viral RNA through a non-specific protein-RNA interaction mechanism. |
| Databáze: | Networked Digital Library of Theses & Dissertations |
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