title: Tetravalent single-chain avidin: from subunits to protein domains via circularly permuted avidins.
abstract: scAvd (single-chain avidin, where two dcAvd are joined in a single polypeptide chain), having four biotin-binding domains, was constructed by fusion of topologically modified avidin units. scAvd showed similar biotin binding and thermal stability properties as chicken avidin. The DNA construct encoding scAvd contains four circularly permuted avidin domains, plus short linkers connecting the four domains into a single polypeptide chain. In contrast with wild-type avidin, which contains four identical avidin monomers, scAvd enables each one of the four avidin domains to be independently modified by protein engineering. Therefore the scAvd scaffold can be used to construct spatially and stoichiometrically defined pseudotetrameric avidin molecules showing different domain characteristics. In addition, unmodified scAvd could be used as a fusion partner, since it provides a unique non-oligomeric structure, which is fully functional with four high-affinity biotin-binding sites. Furthermore, the subunit-to-domain strategy described in the present study could be applied to other proteins and protein complexes, facilitating the development of sophisticated protein tools for applications in nanotechnology and life sciences.
authors.affiliation: Department of Biological and Environmental Science, NanoScience Center, P.O. Box 35 (YAB), FIN-40014 University of Jyväskylä, Finland.
authors.forename: Henri R
authors.initials: HR
authors.lastname: Nordlund
authors.forename: Vesa P
authors.initials: VP
authors.lastname: Hytönen
authors.forename: Jarno
authors.initials: J
authors.lastname: Hörhä
authors.forename: Juha A E
authors.initials: JA
authors.lastname: Määttä
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: White
authors.forename: Katrin
authors.initials: K
authors.lastname: Halling
authors.forename: Eevaleena J
authors.initials: EJ
authors.lastname: Porkka
authors.forename: J Peter
authors.initials: JP
authors.lastname: Slotte
authors.forename: Olli H
authors.initials: OH
authors.lastname: Laitinen
authors.forename: Markku S
authors.initials: MS
authors.lastname: Kulomaa
mesh_terms: D000818:Animals; D001360:Avidin / Q000737:chemistry* / Q000378:metabolism*; D001665:Binding Sites; D002478:Cells, Cultured; D002645:Chickens; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D015202:Protein Engineering / Q000379:methods*; D017434:Protein Structure, Tertiary; D021122:Protein Subunits
chemical_list: D021122:Protein Subunits; D001360:Avidin