title: Probing Ca2+-binding capability of viral proteins with the EF-hand motif by grafting approach.
abstract: Ca(2+) is implicated in almost every step of the life cycle of viruses, including virus entry into host cells, virus replication, virion assembly, maturation, and release. However, due to the lack of prediction algorithms and rigorous validation methods, only limited cases of viral Ca(2+)-binding sites are reported. Here, we introduce a method to predict continuous EF-hand or EF-hand-like motifs in the viral genomes based on their primary sequences. We then introduce a grafting approach, and the use of luminescence resonance energy transfer and Ca(2+) competition assay for experimental verification of predicted Ca(2+)-binding sites. This protocol will be valuable for the prediction and identification of unknown Ca(2+)-binding sites in virus.
authors.affiliation: Department of Chemistry, Georgia State University, Atlanta, GA, USA.
authors.forename: Yubin
authors.initials: Y
authors.lastname: Zhou
authors.forename: Shenghui
authors.initials: S
authors.lastname: Xue
authors.forename: Yanyi
authors.initials: Y
authors.lastname: Chen
authors.forename: Jenny J
authors.initials: JJ
authors.lastname: Yang
mesh_terms: D001665:Binding Sites; D002118:Calcium / Q000378:metabolism*; D019295:Computational Biology / Q000379:methods*; D020832:EF Hand Motifs; D016679:Genome, Viral / Q000235:genetics; D008163:Luminescent Measurements; D008958:Models, Molecular; D010363:Pattern Recognition, Automated; D011485:Protein Binding; D015202:Protein Engineering; D017434:Protein Structure, Tertiary; D014035:Togaviridae / Q000235:genetics; D014764:Viral Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D014764:Viral Proteins; D002118:Calcium