UV resonance Raman study of streptavidin binding of biotin and 2-iminobiotin: comparison with avidin.
UV resonance Raman (UVRR) spectroscopy is used to study the binding of biotin and 2-iminobiotin by streptavidin, and the results are compared to those previously obtained from the avidin-biotin complex and new data from the avidin-2-iminobiotin complex. UVRR difference spectroscopy using 244-nm excitation reveals changes to the tyrosine (Tyr) and tryptophan (Trp) residues of both proteins upon complex formation. Avidin has four Trp and only one Tyr residue, while streptavidin has eight Trp and six Tyr residues. The spectral changes observed in streptavidin upon the addition of biotin are similar to those observed for avidin. However, the intensity enhancements observed for the streptavidin Trp Raman bands are less than those observed with avidin. The changes observed in the streptavidin Tyr bands are similar to those observed for avidin and are assigned exclusively to the binding site Tyr 43 residue. The Trp and Tyr band changes are due to the exclusion of water and addition of biotin, resulting in a more hydrophobic environment for the binding site residues. The addition of 2-iminobiotin results in spectral changes to both the streptavidin and avidin Trp bands that are very similar to those observed upon the addition of biotin in each protein. The changes to the Tyr bands are very different than those observed with the addition of biotin, and similar spectral changes are observed in both streptavidin and avidin. This is attributable to hydrogen bond changes to the binding site Tyr residue in each protein, and the similar Tyr difference features in both proteins supports the exclusive assignment of the streptavidin Tyr difference features to the binding site Tyr 43.
Clarkson
J
J
Department of Physics and Astronomy, Leeds University, Leeds LS2 9JT, England, United Kingdom. phyjcl@phys-irc.novell.leeds.ac.uk
Batchelder
D N
DN
Smith
D A
DA
D001360:Avidin / Q000737:chemistry*; D001665:Binding Sites; D001710:Biotin / Q000031:analogs & derivatives* / Q000737:chemistry*; D046911:Macromolecular Substances; D015394:Molecular Structure; D011485:Protein Binding; D013059:Spectrum Analysis, Raman; D019809:Streptavidin / Q000737:chemistry*; D014364:Tryptophan / Q000737:chemistry; D014443:Tyrosine / Q000737:chemistry
D046911:Macromolecular Substances; D001360:Avidin; D014443:Tyrosine; D001710:Biotin; D014364:Tryptophan; D019809:Streptavidin; C028072:2-iminobiotin