Introduction of histidine residues into avidin subunit interfaces allows pH‐dependent regulation of quaternary structure and biotin binding

FEBS Letters

Uotila, Sanna T.H²  Kulomaa, Markku S³  Niskanen, Einari A³  Savolainen, Janne¹  Hytönen, Vesa P³  Laitinen, Olli H²  Porkka, Eevaleena³  Nordlund, Henri R³ 

[1] Department of Chemistry, P.O. Box 35, FIN-40014 University of Jyväskylä, Finland;A.I. Virtanen Institute, University of Kuopio, FIN-70211 Kuopio, Finland;Department of Biological and Environmental Science, P.O. Box 35, FIN-40014 University of Jyväskylä, Finland

关键词: Avidin;    pH dependence;    Protein engineering;    FCS;    fluorescence correlation spectroscopy;    wt;    wild-type;    Avm;    avidin mutant;   

DOI  :  10.1016/S0014-5793(03)01302-4

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

In order to turn the subunit association and biotin binding of avidin into pH-sensitive phenomena, we have replaced individually three amino acid residues in avidin (Met96, Val115 and Ile117) with histidines in the 1–3 interface, and in combination with a histidine conversion in the 1–2 interface (Trp110). The single replacements Met96His and Val115His in the 1–3 interface were found to have a clear effect on the quaternary structure of avidin, since subunit associations of these mutants became pH-dependent. The histidine replacement in the 1–2 interface affected the biotin-binding properties of the mutants, in particular reversibility of binding and protein–ligand complex formation were pH-sensitive, as measured by IAsys biosensor and fluorescence correlation spectroscopy, respectively. The possibility of regulating the quaternary structure and function of avidin in a controlled and predictable manner, due to introduced interface histidines, will expand even further the range and versatility of the avidin–biotin technology.

【 授权许可】

Unknown