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Vaccinia virus I7L core protein proteinase

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dc.contributor.advisor Hruby, Dennis E.
dc.creator Byrd, Chelsea M.
dc.date.accessioned 2012-06-18T18:14:54Z
dc.date.available 2012-06-18T18:14:54Z
dc.date.copyright 2005-04-08
dc.date.issued 2005-04-08
dc.identifier.uri http://hdl.handle.net/1957/29927
dc.description Graduation date: 2005 en_US
dc.description.abstract Vaccinia virus (VV) is a large double-stranded DNA virus that is a prototypic member of the orthopoxvirus family. Previous works has showed that three of the major structural proteins found within the mature VV virion core 4a, 4b, and 25K are produced from higher molecular weight precursors at late times during infection and processed via a common morphogenic cleavage pathway that is intimately linked with virion assembly and maturation. The enzyme that carries out these cleavage reactions is unknown. A transient expression assay was used to demonstrate that the 17L gene product and its encoded cysteine proteinase activity is responsible for cleavage of each of the three major core protein precursors. Cleavage was demonstrated to occur at the authentic Ala-Gly-Xaa cleavage sites and require active enzyme. A truncated 17L protein lost the ability to cleave the core protein precursors. A conditional-lethal recombinant virus was constructed in which the expression of the 17L gene is under the control of the tetracycline operator/repressor system. In the absence of 17L expression, processing of the major VV core proteins is inhibited and electron microscopy revealed defects in virion morphogenesis prior to complete core condensation. Plasmid-borne 17L is capable of rescuing the growth of this virus. A structural model of 17L was developed and a unique chemical library was assayed for both cell toxicity and the ability to inhibit the growth of VV in tissue culture cells. A novel class of inhibitors was discovered that is capable of inhibiting VV. An in-vitro cleavage assay was developed to further characterize the activity of 17L. This assay is based on producing the major core protein precursors in a coupled transcription and translation assay and then mixing them with 17L enzyme extracts. Using this assay, 17L is shown to be capable of cleavage of each substrate. 17L is further characterized as a cysteine proteinase due to the inhibitory effects of known cysteine proteinase inhibitors such as NEM and iodoacetic acid, as well as through the use of specific small molecule inhibitors in this in-vitro assay. en_US
dc.language.iso en_US en_US
dc.subject.lcsh Vaccinia en_US
dc.subject.lcsh Viral proteinases en_US
dc.subject.lcsh DNA viruses -- Genetics en_US
dc.title Vaccinia virus I7L core protein proteinase en_US
dc.type Thesis/Dissertation en_US
dc.degree.name Doctor of Philosophy (Ph. D.) in Molecular and Cellular Biology en_US
dc.degree.level Doctoral en_US
dc.degree.discipline Science en_US
dc.degree.grantor Oregon State University en_US
dc.contributor.committeemember Arp, Dan
dc.contributor.committeemember Rockey, Dan
dc.contributor.committeemember Ream, Walt
dc.contributor.committeemember Bermudez, Luiz
dc.description.digitization File scanned at 300 ppi (Monochrome, 256 Grayscale, 24-bit Color) using Capture Perfect 3.0.82 on a Canon DR-9080C in PDF format. CVista PdfCompressor 4.0 was used for pdf compression and textual OCR. en_US
dc.description.peerreview no en_us


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