Grant: Research Tools and Instruments

Proteins are biological macromolecules that regulate all life and death processes. This extraordinary diversity of function is determined by the amino acid sequence and environment that, in turn, dictate secondary, tertiary and quaternary structure configurations of the protein. For myriad proteins, folding into highly complex three-dimensional arrangements is required for proper catalytic functions or interactions with other proteins, molecules or ions. Despite the many millions of different proteins that exist in nature, structures are dominated by recurring motifs that include the alpha-helix, beta-strand and random coil conformations. Circular dichroism (CD) spectropolarimetry is the gold standard method for unambiguously measuring the presence of these regular structures, as signals in the far-ultraviolet (UV) region result in similarly shaped spectra for the same secondary structures even when formed by proteins with negligible sequence similarity. Further, the orientation of aromatic residues in non-polar environments not only provides unique CD signals in the near-UV region but also endows proteins with predictable and monitorable intrinsic fluorescent properties; thus, near-UV CD spectropolarimetry and fluorescence are sensitive probes of tertiary and quaternary structures. This multi-applicant proposal urgently requests funds to purchase a CD spectropolarimeter with fluorescence capabilities to be installed in the multi-user, centralized Biomolecular Core Facility (BioCORE) at Western University. The current CD instrument in the BioCORE is ~25 years old and is plagued by frequent breakdowns and obsolete parts, leaving the instrument inoperative for long stretches. Given that CD spectrometry provides fundamental data on the folds of newly expressed proteins, optimal solution conditions for NMR or crystallization, protein stability and structural changes due to binding of ligands or other proteins, modernization with a state-of-the-art CD instrument will not only increase efficiency but also evade the current and looming productivity loss due to catastrophic instrument failure. In addition, the decreased noise, enhanced stability, increased wavelength accuracy and new fluorescence capabilities will make the requested modern instrument more versatile, higher throughput and open new research directions for a large number of NSERC-funded researchers and HQP. A modern CD instrument is a cornerstone resource necessary for continued success of NSERC-funded research programs and the BioCORE facility at Western. The BioCORE is a technology hub for biomolecular research at Western that has supported 100s of researchers and 1000s of trainees over the past 25 years. The centralized location of the facility and professional staff will keep the instrument optimally maintained and managed while offering accessible, expert training for HQP and providing skills needed for careers in academia, government and industry.