title: Theoretical and experimental studies of biotin analogues that bind almost as tightly to streptavidin as biotin.
abstract: We have used a newly developed qualitative computational approach, PROFEC (Pictorial Representation of Free Energy Changes), to visualize the areas of the ligand biotin where modifications of its structure might lead to tighter binding to the protein streptavidin. The PROFEC analysis, which includes protein flexibility and ligand solvation/desolvation, led to the suggestion that the pro-9R hydrogen atom of biotin, which is in alpha-position to the CO(2)(-) group, might be changed to a larger group and lead to better binding with streptavidin and avidin. Free energy calculations supported this suggestion and predicted that the methyl analogue should bind approximately 3 kcal/mol more tightly to streptavidin, with this difference coming exclusively from the relative desolvation free energy of the ligand. The PROFEC analysis further suggested little or no improvement for changing the pro-9S hydrogen atom to a methyl group, and great reduction in changing the ureido N-H groups to N-CH(3). Stimulated by these results, we synthesized 9R-methylbiotin and 9S-methylbiotin, and their binding free energies and enthalpies were measured for interaction with streptavidin and avidin, respectively. In contrast to the calculated results, experiments found both 9-methylbiotin isomers to bind more weakly to streptavidin than biotin. The calculated preference for the binding of the 9R- over the 9S-stereoisomer was observed. In addition, 9-methylbiotin is considerably less soluble in water than biotin, as predicted by the calculation, and the 9R isomer is, to our knowledge, thus far the tightest binding analogue of biotin to streptavidin. Subsequently, X-ray structures of the complexes between streptavidin and both 9R- and 9S-methylbiotin were determined, and the structures were consistent with those used in the free energy calculations. Thus, the reason for the discrepancy between the calculated and experimental binding free energy does not lie in unusual binding modes for the 9-methylbiotins.
authors.affiliation: Department of Pharmaceutical Chemistry, University of California, San Francisco 94143, USA. richard.dixon@abbott.com
authors.forename: Richard W
authors.initials: RW
authors.lastname: Dixon
authors.forename: Randall J
authors.initials: RJ
authors.lastname: Radmer
authors.forename: Bernd
authors.initials: B
authors.lastname: Kuhn
authors.forename: Peter A
authors.initials: PA
authors.lastname: Kollman
authors.forename: Jaemoon
authors.initials: J
authors.lastname: Yang
authors.forename: Cesar
authors.initials: C
authors.lastname: Raposo
authors.forename: Craig S
authors.initials: CS
authors.lastname: Wilcox
authors.forename: Lisa A
authors.initials: LA
authors.lastname: Klumb
authors.forename: Patrick S
authors.initials: PS
authors.lastname: Stayton
authors.forename: Craig
authors.initials: C
authors.lastname: Behnke
authors.forename: Isolde
authors.initials: I
authors.lastname: Le Trong
authors.forename: Ronald
authors.initials: R
authors.lastname: Stenkamp
mesh_terms: D000465:Algorithms; D001360:Avidin / Q000737:chemistry*; D001665:Binding Sites; D001710:Biotin / Q000031:analogs & derivatives* / Q000737:chemistry*; D002384:Catalysis; D002851:Chromatography, High Pressure Liquid; D018360:Crystallography, X-Ray; D009682:Magnetic Resonance Spectroscopy; D008962:Models, Theoretical; D008968:Molecular Conformation; D015394:Molecular Structure; D013237:Stereoisomerism; D019809:Streptavidin / Q000737:chemistry*; D013329:Structure-Activity Relationship
chemical_list: D001360:Avidin; D001710:Biotin; D019809:Streptavidin