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- 1. Curriculum Vitae Michael B. Tropak, PhD 731 Milverton Blvd, Toronto, ON, M4C 1Y2 mb.tropak@utoronto.ca Education 1986-1996 Ph.D., University of Toronto, Dept. of Molecular and Medical Genetics Thesis Title: Myelin Associated Glycoprotein: An I-type lectin. 1979-1983 B.Sc. University of Toronto, Dept. of Medical Microbiology and Parasitology Areas of Expertise Research Experience 2013-present Research Associate, Dr. Jason Maynes, Programme in Molecular Structure and Function, Peter Gilgan Centre for Research and Learning, Sickkids, Toronto 2001-2013 Research Associate, Dr. Don Mahuran, Division of Genetics and Genome Biology, Sickkids, 2006-2011 Research Associate, Dr. Andrew Redington, Dept. of Cardiology and Dr. John Molecular Biology Protein Biochemistry Cell Biology CRISPR Knockout –HEK/HeLa,HT1080 Enzyme kinetics (Glycosidase/transferase) activity DNA transfection (liposomal, Nucleofection), established cell lines (HEK, HeLA, C2C12,HL1, SHSY5Y) CRISPR Knockin – luciferase tagging Enzyme-based assay development Primary cell culture, Oligodendrocyte, Dorsal root ganglion, cerebellar neurons(rat) Piggybac/FLP recombinase transgenic Cell lines Thermofluorescence denaturation shift assay Lentiviral production (HEK293F), infection (C2C12, IPSC CM, HL1) Mutagenesis: PCR/QuikChange/Q5 Drug screening campaigns using Assays described above Immunofluorescent staining endogenous/ Recombinant tagged antigens Cloning: Gateway, Type IIS, Gibson Assembly, PCR/TOPO/TA Protein production (+/-N15 ) purification from E.coli, Pichia, HEK293F Nickel,GST, proteinA,Myc affinity tags Epifluorescence (deconvolution) Confocal imaging, spinning, scanning,SIM PCR: Genomic, Realtime Crystallization Screening High Content Imaging (Cellomics), Image Analysis CellProfiler, ImageJ, Volocity immunoprecipitation and sample prep (+/- isotopic label) for LC/MS proteomics subcellular fractionation of lysosomes and mitochondria using dextran/Ab coated magnetic beads or via sucrose density fractionation Sample Preparation for targeted MS-based metabolomics Cell-based drug screening campaigns (enzyme, luminescent promoter reporters) Gel electrophoresis (1D/2D), Westerns Cell-based Impedance Spectroscopy (ACEA/Excelligence)
- 2. Callahan, Division of Genetics and Genome Biology, Sickkids, 1996-2000 Postdoctoral Research Fellow, Dr. James M. Rini, Dept. Biochemistry, University of Toronto. 1986 -1996 PhD, Graduate Studies, Dept. Molecular and Medical Genetics; University of Toronto. Supervisor: Dr. John C. Roder 1983 – 1986 Technician. Dept. of Genetics. Hospital for Sick Children. Supervisor: Dr. Roy Gravel. Grants (written as Co-investigator): 2005-2007 NIH: High Throughput Cell based screen for compounds enhancing glucocerebrosidase activity in patient cells PI: D. Mahuran Co-Investigator: M.B. Tropak 2007-2012 CIHR Team Grant: CIHR Team in lysosomal storage disease pharmaco- therapeutics. Team Leaders: D. Mahuran (West), S. Withers (East) Co-PI: JTR Clarke, EB Brown, C. Lingwood, M. James, L. Clarke, M. Cuifolini Co- Investigator: M.B. Tropak 2010--2013 APBDRF: Identifying pharmacological chaperones for Adult Polyglucosan Body Disease PI: D. Mahuran, Co-Investigator: M.B. Tropak Grants (Written but requested salary precludes co-investigator status): 2010-2015 CIHR: Emerging Team Grant in Arrhythmogenic Right Ventricular Cardiomyopathy Team Leader: Robert Hamilton Co-PI/Investigators: S. Scherer, D. Mahuran, I Stagljar, G. Wilson, C. Ackerly, N. Kumar, A. Gramolini, M.B. Tropak: 2012-2013 CIHR Proof of Principle Grant 1: Treatment of Parkinson Disease Progression with GCase Activity Enhancers PI: D. Mahuran Co-PI/Investigators: D. Mount and A. Tandon (Univerity of Toronto) Industrial Partner: Exsar Inc. Teaching Experience
- 3. 2010-present Lecturer, Waterloo University, Dept of Biology, year undergraduate course in Glycobiology, Course Coordinators – Dr. David Rose and Dr. Warren Wakarchuk. Two 2 hr lectures on Glycosphingolipidoses and Congenital Disorders of Glycosylation 1999-2000 Lecturer, Ryerson University, Dept. of Chemistry and Biology, 2nd year undergraduate course in Biochemistry and Microbiology. Department Head -Dr. Deborah Foster. 1999-2000 Lecturer, University of Toronto, Faculty of Medicine, Botany and Zoology 3rd year undergraduate course in molecular genetics (JLM349): Course Coordinator-Dr. Sue Varmuza 1993-1996 Teaching assistant, University of Toronto, Molecular Genetics and Biology program, 4th year undergraduate molecular biology laboratory course (MGB430H): Course Coodinator -Dr. Marvin Gold. Awards Ad Hoc Committees Service Excellence Communications Task Group Lead: John Pires (Sept. 2010 – Jan 2010) Purpose: The Fire Starter Campaign -Implementing a culture of service excellence at SickKids over the next three to six months. Connecting Artists at OCAD with Scientists at Sickkids Members: Tropak, Di Carlo- Ancic (Communications at Sickkids), Spicanovic (Dean of Art, OCAD). (April 2013 – Aug 2013) Purpose: Communicating Science through different Art Media: Exploring an Art installation at the Research and Learning Centre in collaboration with OCAD. Publication Summary (1983-2016) Publication Record Summary (2016): h_index: 27, Total 64 publications: Undergrad (1983-84)2, Tech (1986-88)4, PhD (88-96):12 PDF (97-2000); 3, RA (2001-2016) 43 Peer Reviewed Publications (Selection from last 5 years) 1. Tropak, M. B.; Zhang, J.; Yonekawa, S.; Rigat, B. A.; Aulakh, V. S.; Smith, M. R.; Hwang, H. J.; Ciufolini, M. A.; Mahuran, D. J. Pyrimethamine Derivatives: Insight into Binding Mechanism and Improved Enhancement of Mutant beta-N-acetylhexosaminidase Activity. J Med Chem 2015, 58, 4483-93. 2. Tropak, M. B.; Yonekawa, S.; Karumuthil-Melethil, S.; Thompson, P.; Wakarchuk, W.; Gray, S. J.; Walia, J. S.; Mark, B. L.; Mahuran, D. Construction of a hybrid beta-hexosaminidase subunit capable of forming stable homodimers that hydrolyze GM2 ganglioside in vivo. Mol Ther Methods Clin Dev 2016, 3, 15057. 3. Rigat, B. A.; Tropak, M. B.; Buttner, J.; Crushell, E.; Benedict, D.; Callahan, J. W.; Martin, D. R.; Mahuran, D. J. Evaluation of N-nonyl-deoxygalactonojirimycin as a pharmacological chaperone for human GM1 gangliosidosis leads to identification of a feline model suitable for testing enzyme enhancement therapy. Mol
- 4. Genet Metab 2012, 107, 203-12. 4. Goddard-Borger, E. D.; Tropak, M. B.; Yonekawa, S.; Tysoe, C.; Mahuran, D. J.; Withers, S. G. Rapid assembly of a library of lipophilic iminosugars via the thiol-ene reaction yields promising pharmacological chaperones for the treatment of Gaucher disease. J Med Chem 2012, 55, 2737-45. 5. Rempel, B. P.; Tropak, M. B.; Mahuran, D. J.; Withers, S. G. Tailoring the specificity and reactivity of a mechanism-based inactivator of glucocerebrosidase for potential therapeutic applications. Angew Chem Int Ed Engl 2011, 50, 10381-3. Peer Reviewed Publications (Selection from beyond 5 years) 1. Bateman, K. S.; Cherney, M. M.; Mahuran, D. J.; Tropak, M.; James, M. N. Crystal structure of beta- hexosaminidase B in complex with pyrimethamine, a potential pharmacological chaperone. J Med Chem 2011, 54, 1421-9. 2. Clarke, J. T.; Mahuran, D. J.; Sathe, S.; Kolodny, E. H.; Rigat, B. A.; Raiman, J. A.; Tropak, M. B. An open- label Phase I/II clinical trial of pyrimethamine for the treatment of patients affected with chronic GM2 gangliosidosis (Tay-Sachs or Sandhoff variants). Mol Genet Metab 2011, 102, 6-12. 3. Maegawa, G. H.; Tropak, M. B.; Buttner, J. D.; Rigat, B. A.; Fuller, M.; Pandit, D.; Tang, L.; Kornhaber, G. J.; Hamuro, Y.; Clarke, J. T.; Mahuran, D. J. Identification and characterization of ambroxol as an enzyme enhancement agent for Gaucher disease. J Biol Chem 2009, 284, 23502-16. 4. Shimizu, M.; Tropak, M.; Diaz, R. J.; Suto, F.; Surendra, H.; Kuzmin, E.; Li, J.; Gross, G.; Wilson, G. J.; Callahan, J.; Redington, A. N. Transient limb ischaemia remotely preconditions through a humoral mechanism acting directly on the myocardium: evidence suggesting cross-species protection. Clin Sci (Lond) 2009, 117, 191-200. 5. Tropak, M. B.; Kornhaber, G. J.; Rigat, B. A.; Maegawa, G. H.; Buttner, J. D.; Blanchard, J. E.; Murphy, C.; Tuske, S. J.; Coales, S. J.; Hamuro, Y.; Brown, E. D.; Mahuran, D. J. Identification of pharmacological chaperones for Gaucher disease and characterization of their effects on beta-glucocerebrosidase by hydrogen/deuterium exchange mass spectrometry. Chembiochem 2008, 9, 2650-62. 6. Maegawa, G. H.; Tropak, M.; Buttner, J.; Stockley, T.; Kok, F.; Clarke, J. T.; Mahuran, D. J. Pyrimethamine as a potential pharmacological chaperone for late-onset forms of GM2 gangliosidosis. J Biol Chem 2007, 282, 9150-61. 7. Tropak, M. B.; Blanchard, J. E.; Withers, S. G.; Brown, E. D.; Mahuran, D. High-throughput screening for human lysosomal beta-N-Acetyl hexosaminidase inhibitors acting as pharmacological chaperones. Chem Biol 2007, 14, 153-64. 8. Tropak, M. B.; Reid, S. P.; Guiral, M.; Withers, S. G.; Mahuran, D. Pharmacological enhancement of beta- hexosaminidase activity in fibroblasts from adult Tay-Sachs and Sandhoff Patients. J Biol Chem 2004, 279, 13478-87.