Structural and functional characteristics of xenavidin, the first frog avidin from Xenopus tropicalis.
BACKGROUND
Avidins are proteins with extraordinarily high ligand-binding affinity, a property which is used in a wide array of life science applications. Even though useful for biotechnology and nanotechnology, the biological function of avidins is not fully understood. Here we structurally and functionally characterise a novel avidin named xenavidin, which is to our knowledge the first reported avidin from a frog.


RESULTS
Xenavidin was identified from an EST sequence database for Xenopus tropicalis and produced in insect cells using a baculovirus expression system. The recombinant xenavidin was found to be homotetrameric based on gel filtration analysis. Biacore sensor analysis, fluorescently labelled biotin and radioactive biotin were used to evaluate the biotin-binding properties of xenavidin - it binds biotin with high affinity though less tightly than do chicken avidin and bacterial streptavidin. X-ray crystallography revealed structural conservation around the ligand-binding site, while some of the loop regions have a unique design. The location of structural water molecules at the entrance and/or within the ligand-binding site may have a role in determining the characteristic biotin-binding properties of xenavidin.


CONCLUSIONS
The novel data reported here provide information about the biochemically and structurally important determinants of biotin binding. This information may facilitate the discovery of novel tools for biotechnology.
Institute of Medical Technology, Biokatu 6, FI-33014 University of Tampere and Tampere University Hospital, Tampere, Finland. juha.maatta@uta.fi
Juha A E
JA
Määttä
Satu H
SH
Helppolainen
Vesa P
VP
Hytönen
Mark S
MS
Johnson
Markku S
MS
Kulomaa
Tomi T
TT
Airenne
Henri R
HR
Nordlund
D000595:Amino Acid Sequence; D000818:Animals; D001360:Avidin / Q000737:chemistry* / Q000378:metabolism; D001665:Binding Sites; D001710:Biotin / Q000378:metabolism; D018360:Crystallography, X-Ray; D008024:Ligands; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014981:Xenopus
D008024:Ligands; D011994:Recombinant Proteins; D001360:Avidin; D001710:Biotin