Discipline: Biological Sciences
Subcategory: Biochemistry (not Cell and Molecular Biology and Genetics)
Joshua Haynes - Fisk University
Co-Author(s): Ben Gilston and Walter Chazin, Vanderbilt University Dhara Patel, Velia Garcia, and Steven Damo, Fisk University
Calprotectin (CP) is a member of the S100 class of the EF-hand family of calcium binding proteins. It is a heterodimer comprised of S100A8 and S100A9 subunits. In addition to binding calcium, CP can bind a single manganese (II) ion (Mn) at the heterodimer interface. Mn sequestration by CP is important for the innate immune response to bacterial pathogens. CP binds Mn with high affinity to starve pathogens of this essential metal, a process termed nutritional immunity. The binding of Mn is achieved by a hexahistidine motif that is unique to all S100 proteins. The x-ray crystal structure of Mn-CP reveals that the quaternary structure of the complex is a dimer of heterodimers. The objective of this work is to quantitatively characterize how Mn modulates the quaternary structure of CP. Native page experiments indicate that apo CP runs as a single band consistent with heterodimeric structure. The addition of calcium causes several oligomers of CP to form and these oligomers are similar to manganese and calcium bound CP. Dynamic light scattering studies show that apo CP is a monodisperse heterodimer, however metal bound CP exhibits high polydispersity. Together, these experiments represent an important first step toward characterizing the solution structure of Mn-CP and this approach will be broadly applicable to other members of the S100 class of proteins. Future experiments will focus on determining the molecular mass and radii of CP oligomers to enable detailed structural modeling.
Funder Acknowledgement(s): This work was supported by the NSF Award #s 1547757, 1500320, and 1400969.
Faculty Advisor: Steven Damo, email@example.com
Role: Site directed mutagenesis, protein growths, affinity chromatography , size exclusion chromatography, protein crystallization, X-ray crystallography techniques, growth experiments.