CRYSTALLOGRAPHIC ANALYSES OF UNLIGANDED GIANT ANNELID RESPIRATORY PROTEINS

这个子项目是众多研究子项目之一

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Extracellular annelid respiratory proteins assemble into large complexes (up to 3,600 kDa), which allows substantial concentration of oxygen binding capacity into a single particle. Befitting their large size, these proteins exhibit highly cooperative oxygen binding properties that are dependent upon the interplay of divalent cations and protons. The goal of this proposal is to investigate the structural basis for the strong allosteric properties of these large respiratory proteins. Our subjects are three distinct, but related, cooperative annelid complexes. These include two 3,600 kDa assemblages, the erythrocruorins from Lumbricus terrestris and Arenicola marina and a 400 kDa hemoglobin found in the hydrothermal vent tubeworm, Riftia pachyptila. We have determined crystal structures of each complex with bound ligand (either oxygen or carbon monoxide), using diffraction data collected from BioCARS beamline 14-BMC. The crystal structures of both 3,600 kDa erythrocruorins reveal 180 polypeptide chains that form hexagonal bilayer structures, but with altered arrangements of the hexagonal layers stemming from differences in heterotrimeric coiled-coil domains near the center of each complex. The 400 kDa Riftia complex structure is a hollow D3 symmetric sphere of 24 subunits, with each half arranged similarly to dodecamer complexes found in the erythrocruorins (Flores et al., PNAS 102, 2713-2718, 2005). There are currently no unliganded crystal structures available for any members of this family of respiratory proteins. We have recently grown crystals of each of these proteins in the absence of ligand. Crystal structures of these proteins will allow us to understand the ligand-linked structural transitions that mediate cooperativity and their comparison will reveal the degree to which each similar, but distinct, quaternary arrangement impacts the range of ligand-linked transitions and how this correlates with the functional properties.