个人简介

Ph.D. University of Toronto 2001 B.Sc. University of Western Ontario 1995

研究领域

Structural Biology

Research in the laboratory is focused primarily on the sulfonylurea receptor (SUR) protein family. SUR proteins are members of the ATP-binding cassette (ABC) superfamily of transporters, which are integral membrane proteins found in all species. Unlike most ABC proteins, the SUR proteins do not possess any transporter activity, but form the regulatory subunits in ATP-sensitive potassium (K-ATP) channels. Four copies of the SUR proteins surround four copies of the pore-forming Kir6.x (Kir6.1 or Kir6.2) proteins to form K-ATP channels. Different combinations of SUR and Kir6.x proteins combine to form K-ATP channels in various tissues. ATP binding at the Kir6.x proteins results in K-ATP channel inhibition whereas MgATP binding and hydrolysis at the SUR NBDs results in channel opening. NBD1 and NBD2 of SUR2A are also sites of phosphorylation by protein kinase A, which results in results in KATP channel activation. SUR-mediated regulation of KATP channels is critical for cardiovascular and pancreatic function. Over 150 different mutations that cause diseases related to insulin production have been identified in the various regions of SUR1, including in the NBDs. Mutations in the NBDs of SUR2A cause dilated cardiomyopathy, atrial fibrillation, and increased risk of myocardial infarction. Therefore, studies of the SUR NBDs are critical to understanding the mechanism of K-ATP channel regulation and its dysfunction in diseases. We have adopted an interdisciplinary approach to studies of the SUR NBDs. We use a variety of biophysical tools (NMR spectroscopy, CD spectroscopy, fluorescence spectroscopy), biochemical methods (limited proteolysis, nucleotide binding and hydrolysis assays), and bioinformatics analysis of NBD and ABC structure in the research. Our research has already provided insights into the structural domain boundaries and regulatory regions of NBD1 from SUR2A and SUR1 (De Araujo … Kanelis, 2011). We have also examined the effects of the disease-causing mutation V730I (De Araujo, Lopez-Alonso, Kanelis, manuscript in preparation) and phosphorylation (De Araujo, Alvarez, Kanelis, manuscript in preparation) on the structure and dynamics of SUR2A. Further, we have demonstrated that SUR2A NBD1 is a site of interaction of the KATP channel opener (KCO) drug pinacidil, and that drug-binding increases the affinity of SUR2A NBD1 for nucleotide (López-Alonso, De Araujo, Kanelis, 2013), which has important implications for the mechanism of action of this widely studied drug. Currently, we are studying the interaction of SUR2A NBD1 with novel compounds. We are also studying the interaction of SUR2A NBD1 with different regions of SUR2A, and are extending these studies to SUR1. Further, we are currently running experiments to assign the NMR spectra of SUR2A NBD1 so that our interaction, mutation, and phosphorylation studies may be analyzed at the level of individual residues.

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Successful development and use of a thermodynamic stability screen for optimizing the yield of nucleotide binding domains.de Araujo ED, Kanelis V.Protein Expr. Purif. 2014 Nov;103:38-47 pubmed HNH proteins are a widespread component of phage DNA packaging machines.Kala S, Cumby N, Sadowski PD, Hyder BZ, Kanelis V, Davidson AR, Maxwell KL.Proc. Natl. Acad. Sci. U.S.A. 2014 Apr;111(16):6022-7 pubmed NMR and fluorescence studies of drug binding to the first nucleotide binding domain of SUR2A.López-Alonso JP, de Araujo ED, Kanelis V.Biochemistry 2012 Nov;51(45):9211-22 pubmed The protein gp74 from the bacteriophage HK97 functions as a HNH endonuclease.Moodley S, Maxwell KL, Kanelis V.Protein Sci. 2012 Jun;21(6):809-18 pubmed The first nucleotide binding domain of the sulfonylurea receptor 2A contains regulatory elements and is folded and functions as an independent module.de Araujo ED, Ikeda LK, Tzvetkova S, Kanelis V.Biochemistry 2011 Aug;50(31):6655-66 pubmed NMR spectroscopy to study the dynamics and interactions of CFTR.Kanelis V, Chong PA, Forman-Kay JD.Methods Mol. Biol. 2011;741:377-403 pubmed The solution structure of the C-terminal Ig-like domain of the bacteriophage λ tail tube protein.Pell LG, Gasmi-Seabrook GM, Morais M, Neudecker P, Kanelis V, Bona D, Donaldson LW, Edwards AM, Howell PL, Davidson AR, Maxwell KL.J. Mol. Biol. 2010 Oct;403(3):468-79 pubmed NMR evidence for differential phosphorylation-dependent interactions in WT and DeltaF508 CFTR.Kanelis V, Hudson RP, Thibodeau PH, Thomas PJ, Forman-Kay JD.EMBO J. 2010 Jan;29(1):263-77 pubmed The phage lambda major tail protein structure reveals a common evolution for long-tailed phages and the type VI bacterial secretion system.Pell LG, Kanelis V, Donaldson LW, Howell PL, Davidson AR.Proc. Natl. Acad. Sci. U.S.A. 2009 Mar;106(11):4160-5 pubmed Regulation of Nedd4-2 self-ubiquitination and stability by a PY motif located within its HECT-domain.Bruce MC, Kanelis V, Fouladkou F, Debonneville A, Staub O, Rotin D.Biochem. J. 2008 Oct;415(1):155-63 pubmedCFTR regulatory region interacts with NBD1 predominantly via multiple transient helices.Baker JM, Hudson RP, Kanelis V, Choy WY, Thibodeau PH, Thomas PJ, Forman-Kay JD.Nat. Struct. Mol. Biol. 2007 Aug;14(8):738-45 pubmed Nuclear magnetic resonance (NMR) solution structure, dynamics, and binding properties of the kringle IV type 8 module of apolipoprotein(a).Chitayat S, Kanelis V, Koschinsky ML, Smith SP.Biochemistry 2007 Feb;46(7):1732-42 pubmed