Biochemical Characterization of Middle East Respiratory Syndrome Coronavirus Spike Protein Proteolytic Processing.
Identifieur interne : 000293 ( PubMed/Checkpoint ); précédent : 000292; suivant : 000294Biochemical Characterization of Middle East Respiratory Syndrome Coronavirus Spike Protein Proteolytic Processing.
Auteurs : Gary R. Whittaker [États-Unis] ; Jean K. Millet [États-Unis]Source :
- Methods in molecular biology (Clifton, N.J.) [ 1940-6029 ] ; 2020.
Abstract
The coronavirus spike envelope glycoprotein is an essential viral component that mediates virus entry events. Biochemical assessment of the spike protein is critical for understanding structure-function relationships and the roles of the protein in the viral life cycle. Coronavirus spike proteins are typically proteolytically processed and activated by host cell enzymes such as trypsin-like proteases, cathepsins, or proprotein-convertases. Analysis of coronavirus spike proteins by western blot allows the visualization and assessment of proteolytic processing by endogenous or exogenous proteases. Here, we present a method based on western blot analysis to investigate spike protein proteolytic cleavage by transient transfection of HEK-293 T cells allowing expression of the spike protein of the highly pathogenic Middle East respiratory syndrome coronavirus in the presence or absence of a cellular trypsin-like transmembrane serine protease, matriptase. Such analysis enables the characterization of cleavage patterns produced by a host protease on a coronavirus spike glycoprotein.
DOI: 10.1007/978-1-0716-0211-9_3
PubMed: 31883085
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Whittaker</name><affiliation wicri:level="4"><nlm:affiliation>Department of Microbiology and Immunology, College of Veterinary Medicine, Cornell University, Ithaca, NY, USA.</nlm:affiliation><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Microbiology and Immunology, College of Veterinary Medicine, Cornell University, Ithaca, NY</wicri:regionArea><placeName><region type="state">État de New York</region><settlement type="city">Ithaca (New York)</settlement></placeName><orgName type="university">Université Cornell</orgName></affiliation></author><author><name sortKey="Millet, Jean K" sort="Millet, Jean K" uniqKey="Millet J" first="Jean K" last="Millet">Jean K. Millet</name><affiliation wicri:level="4"><nlm:affiliation>Department of Microbiology and Immunology, College of Veterinary Medicine, Cornell University, Ithaca, NY, USA. jean.millet@inra.fr.</nlm:affiliation><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Microbiology and Immunology, College of Veterinary Medicine, Cornell University, Ithaca, NY</wicri:regionArea><placeName><region type="state">État de New York</region><settlement type="city">Ithaca (New York)</settlement></placeName><orgName type="university">Université Cornell</orgName></affiliation></author></analytic><series><title level="j">Methods in molecular biology (Clifton, N.J.)</title><idno type="eISSN">1940-6029</idno><imprint><date when="2020" type="published">2020</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">The coronavirus spike envelope glycoprotein is an essential viral component that mediates virus entry events. Biochemical assessment of the spike protein is critical for understanding structure-function relationships and the roles of the protein in the viral life cycle. Coronavirus spike proteins are typically proteolytically processed and activated by host cell enzymes such as trypsin-like proteases, cathepsins, or proprotein-convertases. Analysis of coronavirus spike proteins by western blot allows the visualization and assessment of proteolytic processing by endogenous or exogenous proteases. Here, we present a method based on western blot analysis to investigate spike protein proteolytic cleavage by transient transfection of HEK-293 T cells allowing expression of the spike protein of the highly pathogenic Middle East respiratory syndrome coronavirus in the presence or absence of a cellular trypsin-like transmembrane serine protease, matriptase. Such analysis enables the characterization of cleavage patterns produced by a host protease on a coronavirus spike glycoprotein.</div></front></TEI><pubmed><MedlineCitation Status="In-Process" Owner="NLM"><PMID Version="1">31883085</PMID><DateRevised><Year>2020</Year><Month>04</Month><Day>18</Day></DateRevised><Article PubModel="Print"><Journal><ISSN IssnType="Electronic">1940-6029</ISSN><JournalIssue CitedMedium="Internet"><Volume>2099</Volume><PubDate><Year>2020</Year></PubDate></JournalIssue><Title>Methods in molecular biology (Clifton, N.J.)</Title><ISOAbbreviation>Methods Mol. 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Here, we present a method based on western blot analysis to investigate spike protein proteolytic cleavage by transient transfection of HEK-293 T cells allowing expression of the spike protein of the highly pathogenic Middle East respiratory syndrome coronavirus in the presence or absence of a cellular trypsin-like transmembrane serine protease, matriptase. Such analysis enables the characterization of cleavage patterns produced by a host protease on a coronavirus spike glycoprotein.</AbstractText></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Whittaker</LastName><ForeName>Gary R</ForeName><Initials>GR</Initials><AffiliationInfo><Affiliation>Department of Microbiology and Immunology, College of Veterinary Medicine, Cornell University, Ithaca, NY, USA.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Millet</LastName><ForeName>Jean K</ForeName><Initials>JK</Initials><AffiliationInfo><Affiliation>Department of Microbiology and Immunology, College of Veterinary Medicine, Cornell University, Ithaca, NY, USA. jean.millet@inra.fr.</Affiliation></AffiliationInfo><AffiliationInfo><Affiliation>Virologie et Immunologie Moléculaires, INRA, Jouy-en-Josas, France. jean.millet@inra.fr.</Affiliation></AffiliationInfo></Author></AuthorList><Language>eng</Language><GrantList CompleteYN="Y"><Grant><GrantID>R01 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