{"id":{"repo_id":"wku-diss","oai_identifier":"oai:digitalcommons.wku.edu:theses-1888"},"canonical_url":"https://search.dev.ndltd.org/etd/wku-diss/oai:digitalcommons.wku.edu:theses-1888","repository":{"repo_id":"wku-diss","name":"Western Kentucky University","base_url":"https://digitalcommons.wku.edu/do/oai/"},"display":{"title":"Site-Directed Mutagenesis of Southern Bean Mosaic Virus (Cowpea Strain): in Search of a Viral Protease","abstract":"The second and largest open reading frame within the southern bean mosaic virus (SBMV) genome encodes a 105 kDa polyprotein. Following translation, the polyprotein is cleaved to liberate various proteins necessary for SBMV replication. The elements within polyproteins of the picornavirus superfamily members have a conserved order: Vpg (viral protein, genome-linked)-protease-replicase. Amino acid sequence homologies indicate that the 105 kDa protein of SBMV contains a replicase very similar to those identified in polyproteins of the picorna-like viruses. The presence of a VPg covalently attached to the 5' end of the SBMV genome further suggests that SBMV may be considered a member of the picornavirus superfamily-Serine 558 within the SBMV polyprotein has been proposed to be a catalytic residue of a serine protease. Site-directed mutagenesis was used to create a mutant with a glycine at this position, and coupled in vitro transcription/translation was used to prepare 3H labeled translation products. SDS-PAGE and fluorography were then used to assay for the presence or absence of polyprotein cleavage. Although site-directed mutagenesis was successful in creating the mutant, a possible deletion that complicated the interpretation of the results was identified.","abstract_html":"The second and largest open reading frame within the southern bean mosaic virus (SBMV) genome encodes a 105 kDa polyprotein. Following translation, the polyprotein is cleaved to liberate various proteins necessary for SBMV replication. The elements within polyproteins of the picornavirus superfamily members have a conserved order: Vpg (viral protein, genome-linked)-protease-replicase. Amino acid sequence homologies indicate that the 105 kDa protein of SBMV contains a replicase very similar to those identified in polyproteins of the picorna-like viruses. The presence of a VPg covalently attached to the 5' end of the SBMV genome further suggests that SBMV may be considered a member of the picornavirus superfamily-Serine 558 within the SBMV polyprotein has been proposed to be a catalytic residue of a serine protease. Site-directed mutagenesis was used to create a mutant with a glycine at this position, and coupled in vitro transcription/translation was used to prepare 3H labeled translation products. SDS-PAGE and fluorography were then used to assay for the presence or absence of polyprotein cleavage. Although site-directed mutagenesis was successful in creating the mutant, a possible deletion that complicated the interpretation of the results was identified.","abstract_has_math":false,"creators":["Simmons, Alan"],"institution":null,"degree_name":"Master of Science","degree_level":null,"degree_discipline":"Department of Biology","degree_department":null,"school":null,"contributors":[],"advisors":[],"committee_chairs":[],"committee_members":[],"year":1995,"date_issued":"1995-01-01T08:00:00Z","date_published":"1995-01-01T08:00:00Z","updated_at":"2026-07-24T06:07:54Z","subjects":["Medical Sciences"],"languages":[],"rights":[],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":"https://digitalcommons.wku.edu/theses/885","outbound_label":"Repository record","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:creator","label":"Author","values":["Simmons, Alan"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:type","label":"Dc Type","values":["Thesis"]},{"key":"thesis:degree_discipline","label":"Discipline","values":["Department of Biology"]},{"key":"thesis:degree_name","label":"Degree Name","values":["Master of Science"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["Medical Sciences"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["https://digitalcommons.wku.edu/theses/885"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["The second and largest open reading frame within the southern bean mosaic virus (SBMV) genome encodes a 105 kDa polyprotein. Following translation, the polyprotein is cleaved to liberate various proteins necessary for SBMV replication. The elements within polyproteins of the picornavirus superfamily members have a conserved order: Vpg (viral protein, genome-linked)-protease-replicase. Amino acid sequence homologies indicate that the 105 kDa protein of SBMV contains a replicase very similar to those identified in polyproteins of the picorna-like viruses. The presence of a VPg covalently attached to the 5' end of the SBMV genome further suggests that SBMV may be considered a member of the picornavirus superfamily-Serine 558 within the SBMV polyprotein has been proposed to be a catalytic residue of a serine protease. Site-directed mutagenesis was used to create a mutant with a glycine at this position, and coupled in vitro transcription/translation was used to prepare 3H labeled translation products. SDS-PAGE and fluorography were then used to assay for the presence or absence of polyprotein cleavage. Although site-directed mutagenesis was successful in creating the mutant, a possible deletion that complicated the interpretation of the results was identified."]},{"key":"dc:title","label":"Title","values":["Site-Directed Mutagenesis of Southern Bean Mosaic Virus (Cowpea Strain): in Search of a Viral Protease"]}]}],"canonical_facts":{"dc:creator":["Simmons, Alan"],"dc:description.abstract":["The second and largest open reading frame within the southern bean mosaic virus (SBMV) genome encodes a 105 kDa polyprotein. Following translation, the polyprotein is cleaved to liberate various proteins necessary for SBMV replication. The elements within polyproteins of the picornavirus superfamily members have a conserved order: Vpg (viral protein, genome-linked)-protease-replicase. Amino acid sequence homologies indicate that the 105 kDa protein of SBMV contains a replicase very similar to those identified in polyproteins of the picorna-like viruses. The presence of a VPg covalently attached to the 5' end of the SBMV genome further suggests that SBMV may be considered a member of the picornavirus superfamily-Serine 558 within the SBMV polyprotein has been proposed to be a catalytic residue of a serine protease. Site-directed mutagenesis was used to create a mutant with a glycine at this position, and coupled in vitro transcription/translation was used to prepare 3H labeled translation products. SDS-PAGE and fluorography were then used to assay for the presence or absence of polyprotein cleavage. Although site-directed mutagenesis was successful in creating the mutant, a possible deletion that complicated the interpretation of the results was identified."],"dc:identifier":["https://digitalcommons.wku.edu/theses/885"],"dc:subject":["Medical Sciences"],"dc:title":["Site-Directed Mutagenesis of Southern Bean Mosaic Virus (Cowpea Strain): in Search of a Viral Protease"],"dc:type":["Thesis"],"thesis:degree_discipline":["Department of Biology"],"thesis:degree_name":["Master of Science"]},"updated_at":"2026-07-24T06:07:54Z"}