In the last year, we have applied our method of computationally accounting for surface site heterogeneity to a study of a model protein immobilized with different chemical strategies and on different sensor surfaces. We found that the most uniform surface site distributions were achieved with linker layers of moderate length. We identified experimental conditions that impose increased mass transport limitations. As an additional result of these studies, we have released an updated version of our distributable SPR data anlaysis software. This now also includes a discrete conformational change model, which allows testing of how well the hypothesis of slow conformational changes after an initial binding step fits experimental multi-phasic surface binding data, in comparison with the alternative explanation of surface site distributions with polydisperse affinity and kinetic constants. This was applied to data from the laboratory of Dr. Birgit Helm. Finally, we have continued the collaboration with the laboratory of Dr. Michael Tarlov at NIST on the recognition of different carbohydrate moieties on glycoproteins by surface-immobilized lectins.
|Zhao, Huaying; Gorshkova, Inna I; Fu, Gregory L et al. (2013) A comparison of binding surfaces for SPR biosensing using an antibody-antigen system and affinity distribution analysis. Methods 59:328-35|
|Zhao, Huaying; Brown, Patrick H; Schuck, Peter (2011) On the distribution of protein refractive index increments. Biophys J 100:2309-17|
|Schuck, Peter; Zhao, Huaying (2010) The role of mass transport limitation and surface heterogeneity in the biophysical characterization of macromolecular binding processes by SPR biosensing. Methods Mol Biol 627:15-54|
|Gondeau, Claire; Corradin, Giampietro; Heitz, Frédéric et al. (2009) The C-terminal domain of Plasmodium falciparum merozoite surface protein 3 self-assembles into alpha-helical coiled coil tetramer. Mol Biochem Parasitol 165:153-61|