Journal
FEBS LETTERS
Volume 555, Issue 3, Pages 449-454Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/S0014-5793(03)01302-4
Keywords
avidin; pH dependence; protein engineering
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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. (C) 2003 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.
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