Biochemistry Seminar - Dr. Voula Kanelis

Dr. Voula Kanelis
Department of Chemical and Physical Sciences, University of Toronto Mississauga

“Biochemical and biophysical studies of the first nucleotide binding domain of SUR2A”

The sulfonylurea receptor 2A (SUR2A) is an ATP-binding cassette (ABC) protein that forms the regulatory subunit of ATP-sensitive potassium (KATP) channels in the heart.  In KATP channels, four SUR2A proteins surround four pore-forming Kir6.2 subunits. Regulation of KATP channel gating is a complex process that involves both the Kir6.2 and SUR2A subunits. ATP binding at the Kir6.2 subunits results in closed KATP channels, whereas ATP binding and hydrolysis at the SUR2A NBDs opens the pore. Additional modes of KATP channel regulation involve phosphorylation by protein kinase A of specific sites within the SUR2A NBDs, with mono-phosphorylation of NBD1 or NBD2 resulting in KATP channel activation and di-phosphorylation further stimulating channel activity. Proper regulation of KATP channels by SUR2A is critical, as mutations in the NBDs that affect ATP hydrolysis and cellular trafficking cause cardiovascular disorders.

We have focused our initial studies of SUR2A on NBD1. Using structure-based sequence alignments, predictions of disordered regions, and biophysical studies, we determined the domain boundaries for SUR2A NBD1 that have enabled, for the first time, NMR studies of NBD1. Surprisingly our alignment and screening data indicate that SUR2A NBD1 contains two putative, previously unidentified, regulatory elements: a large insert within the β-sheet subdomain and a C-terminal extension that are analogous to the RI and RE in CFTR NBD1, and may be present in other members of the C-subfamily of ABC transporters. NMR data on SUR2A NBD1 indicate significant protein dynamics that are altered with ATP binding and phosphorylation and fluorescence data indicate conformational changes in NBD1 with phosphorylation. These data will be presented and discussed in terms of a model that correlates structural changes in SUR2A NBD1 to regulation of KATP channel gating.

Mandatory for Graduate Students