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dc.creatorWu, Xiaofengen_US
dc.date.accessioned2012-06-07T23:19:35Z
dc.date.available2012-06-07T23:19:35Z
dc.date.created2002en_US
dc.date.issued2002
dc.identifier.urihttp://hdl.handle.net/1969.1/ETD-TAMU-2002-THESIS-W92en_US
dc.descriptionDue to the character of the original source materials and the nature of batch digitization, quality control issues may be present in this document. Please report any quality issues you encounter to digital@library.tamu.edu, referencing the URI of the item.en_US
dc.descriptionIncludes bibliographical references (leaves 79-93).en_US
dc.descriptionIssued also on microfiche from Lange Micrographics.en_US
dc.description.abstractAutographa californica nuclear polyhedrosis virus (AcNPV) late genes are transcribed by a viral-encoded RNA polymerase that is composed of four subunits. The LEF-4 subunit has guanylyltransferase and RNA triphosphatase activities which are needed to form the 5' cap structure. But whether AcNPV encodes its own capping methyltransferases (MTase) is yet unknown. The ORF69 protein was identified as a candidate for capping MTases based on its sequence similarity to FtsJ. ORF69 was overexpressed in E.coli and purified to homogeneity. Purified ORF69 had AdoMet binding activity. The pattern of protein expression in vivo and its dependency on DNA replication suggest that ORF69 is a late protein. Primer extension analysis identified the presence of a late promoter motif at the transcription start site, thus further confirming this hypothesis. A mutant virus with the E.coli lacZ gene inserted into the middle of orf69 was constructed and orf69 was shown to be non-essential for virus replication. Single-step growth curves of mutant and wild type viruses showed that the rate of virus replication was not affected by the absence of ORF69. As an approach to the identification of the viral mRNA (guanine-7-) methyltransferase (cap MTase), the enzymatic activity was partially purified from the infected and uninfected cells. Cap MTase exhibited identical purification patterns from both sources on three different columns. Together, these data showed that the AcNPV orf69 gene encodes a late, non-essential protein with the potential to be a MTase. Partial purification of cap MTase from infected cells suggests that AcNPV uses host cap MTase to cap its late transcripts.en_US
dc.format.mediumelectronicen_US
dc.format.mimetypeapplication/pdfen_US
dc.language.isoen_USen_US
dc.publisherTexas A&M Universityen_US
dc.rightsThis thesis was part of a retrospective digitization project authorized by the Texas A&M University Libraries in 2008. Copyright remains vested with the author(s). It is the user's responsibility to secure permission from the copyright holder(s) for re-use of the work beyond the provision of Fair Use.en_US
dc.subjectbiochemistry.en_US
dc.subjectMajor biochemistry.en_US
dc.titlePurification and characterization of ORF69 of Autographa californica nuclear polyhedrosis virusen_US
dc.typeThesisen_US
thesis.degree.disciplinebiochemistryen_US
thesis.degree.nameM.S.en_US
thesis.degree.levelMastersen_US
dc.type.genrethesis
dc.type.materialtexten_US
dc.format.digitalOriginreformatted digitalen_US


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