John Sumida, Ph.D. Chemistry Biophysics/Spectroscopy/ Protein Chemistry E-mail: sumida@bbri.org or jpsumida@comcast.net Cell: 617-877-3002

Summary: I have 10+ years of experience in biophysical characterization of proteins and their biochemistry using a variety of techniques: CD, DSC, TR-FRET, steady state fluorescence, UV/vis spectroscopy, recombinant protein expression, protein purification, FPLC, N-terminal amino acid analysis, , MALDI, chemical cross-linking, and fluorescent labeling of proteins.

Accomplishments:

Established a new Biophysical Core and Cost Facility at the University of Washington. Characterized a new dynamic region in the middle of tropomyosin that is important in muscle

regulation and may have more general implications in muscle related diseases. Presented a platform presentation at the 2008 Biophysical Meeting, entitled “Conserved

Destabilization of Tropomyosin at Asp 137: Functional Significance of Flexibility.” Poster award for best poster at the 2008 Myofilament Meeting Quantitative determination of N-terminal sequences from a mixture of proteolytic fragments. Promotion to Research Associate in 2007. First author JBC paper: “Conserved Asp-137 Imparts Flexibility to Tropomyosin and Affects

Function” First author J. Phys Chem paper: "Contrasting Photoinduced Electron Transfer Properties of

Two Closely Related Rigidly-Linked Porphyrin-Quinone Dyads," First characterization of the photophysical properties of a buckminster fullerene: "Preparation

and Photophysical Studies of Porphyrin-C60 Dyads,"

Professional Work Experience

University of Washington Bioanalytical Pharmacy Core, (2010-current)

Biophysical Core Facility Manager.Responsibilities:

1. act as point of contact for industry and academic scientists interested in performing biophysical measurements within the core.

2. Supervise two facility staff scientists.3. establish accounting procedures that enable use of the facility by both industry and academic users4. manage a budget, generate quarterly reports, hire staff5. create and manage a website for the facility6. consult with users about various biophysical approaches available within the core and assist them in

developing analytical methods suitable for their objectives. The Biophysical Core provides instrumental expertise in DSC, ITC, SPR, and AUC. For more information see the link to the website: http://depts.washington.edu/cidb4bio/index.shtml

7. provide training and workshops to graduate students, post-docs and visiting scientists8. handle all the day to day instrument scheduling, maintenance, and procurement for the lab.9. Facilitate a monthly biophysics user group meeting open to both industry and academic scientists.

Boston University, (2009-current)

Position: Research Assistant

Molecular dynamics simulations of tropomyosin using CHARMM, calculation of dynamic and static persistence lengths of a series of tropomyosin fragments to quantitate flexibility.

Boston Biomedical Research Institute, (2002-2009).

Position: Research Associate.The research objective at BBRI was to address a central and persistent problem in our understanding of how the muscle thin filament is physically able to modulate the regulatory states of the muscle system. The following techniques were used to identify a new flexible region in Tm which affects its conformational dynamics, and regulatory function:

Limited proteolysis, CD, DSC, UV/vis spectroscopy, cosedimentation analyses and ATPase kinetics Time resolved fluorescence (time domain)

This work is published in JBC (see below).

This position at BBRI also involves the review of outside scientific work being considered for publication. Peer review was provided for:

Journal of Molecular Biology, Journal of Biological Chemistry, Proteins

The position at BBRI involves a significant amount of supervision and management of the laboratory’s day to day function. These duties include

supervision of technical assistants training of colleagues and visiting scientists mentorship of student interns on their research projects instructor in institute biophysics

Universities Space Research Assoc. /NASA Marshall Space Flight Center, (1999-2002). Position: Staff Scientist (contractor). The research task at Marshall was to detect and characterize protein nucleation in solution. The following techniques were used to correlate solution distances with the nearest neighbor crystallographic distance:

Fluorescence CD

Education1993 - 1999, Ph.D. Chemistry, Arizona State University, Tempe, AZ

1990 - 1992, B.S. Biochemistry, Seattle University, Seattle, WA

1982 - 1988, B.A. Anthropology, Brown University, Providence, RI

Professional Associations:

Biophysical Society Member since 2001AAAS Member since 2005

Biophysical Expertise steady-state and time-resolved,

(both frequency and time domain) TR-FRET intermolecular

distances and dynamics Stern Volmer quenching to assess

site accessibility. Rotational anisotropy to detect

conformational changes Binding constants Synchronously scanned emission

spectroscopy for the analysis of heterogeneous solutions

domain specific unfolding, determination of thermodynamic

constants, H, G, S, Cp G measurements to assess mutational

effects on thermodynamics and structure Pressure perturbation calorimetry to

determine partial volume of unfolding and thermal expansion coeffiecient

Fluorescence

UV/vis

DSC

Sedimentation velocity using o Interferenceo Absorption o Fluorescence

ion exchange, hydrophobic interaction, mixed

mode (hydroxyapatite) size exclusion chromatographyChromatography

helix unfolding near uv analysis for tertiary

interactions

CD

Standard Biochemical techniques

SDS-PAGE, N-terminal sequence analysis, Chemical modification of proteins, Proteolysis, MALDI-ToF Mass Spec, cosedimentation analyses, bacterial expression and purification of recombinant proteins, FPLC chromatography, IEX, SEC, HIC,

recombinant protein expression in bacterial cells

protein purification coursework in AUC synthetic boundary

measurements dynamic light scattering

AUC

concentration determination,

determination of labeling efficiency,

kinetic enzymatic analysesSPR Kinetics and equilibrium

mode affinity measurements using a Biacrore T100

Important Publications [citations]1. “Conserved Asp-137 Imparts Flexibility to Tropomyosin and Affects Function.” Sumida, J.P., Wu, E., Lehrer,

S.S., J. Biol. Chem., 283, 6728-6734 (2008)

2. “Preparation and preliminary characterization of crystallizing fluorescent derivatives of chicken egg white lysozyme,” J. P. Sumida, E. L. Forsythe, M. L. Pusey, J. Crystal Growth, 232, 308–316 (2001).

3. Pusey, Marc L; Sumida, John P. "Fluorescence of protein crystal nucleation" Proceedings of SPIE vol.4098, Optical Devices and Diagnostics in Materials Science, ed. Andrews, David L.; Asakura, Toshimitsu; Jutamulia, Suganda; Kirk, Wiley P.; Lagally, Max G.; Ravindra B.; Trolinger, James D. September 2000.

4. "Contrasting Photoinduced Electron Transfer Properties of Two Closely Related Rigidly-Linked Porphyrin-Quinone Dyads," J. P. Sumida, P. A. Liddell, A. N. Macpherson, G. R. Seely, A. L. Moore, T. A. Moore and D. Gust, J. Phys. Chem. A, 102, 5512-5519 (1998)

5. “Increasing the yield of photoinduced charge separation through parallel electron transfer pathways,” Maniga, N.I., Sumida, J.P., Stone, S., Moore, A.L., Moore, A.S., Gust, D., J. Porphyrins and Pthalocyanines, 3, 32-34, (1999)

6. "Photoinduced Charge Separation and Charge Recombination to a Triplet State in a Carotene-Porphyrin-Fullerene Triad," P. A. Liddell, D. Kuciauskas, J. P. Sumida, B. Nash, D. Nguyen, A. L. Moore, T. A. Moore and D. Gust, J. Am. Chem. Soc., 119, 1400-1405 (1997).

7. "Preparation and Photophysical Studies of Porphyrin-C60 Dyads," P. A. Liddell, J. P. Sumida, A. N. Macpherson, L. Noss, G. R. Seely, K. N. Clark, A. L. Moore, T. A. Moore and D. Gust, Photochem. Photobiol. 60, 537 - 541 (1994).

8. “Solvent dependence of photoinduced electron transfer in porphyrin dyads,” DeGraziano J.M., Macpherson A.N., Liddell P.A., Noss L., Sumida J.P., Seely G.R., Lewis J.E., Moore A.L., Moore T.A., Gust D., New J. Chem., 20, 839-851, (1996)

Poster Presentations47th annual Biophysical Society Meeting: poster presentation

J. P. Sumida, N. Kirkland, and M. Pusey. Molecular response of chicken egg white lysozyme to buffer, salt concentration and pH.

49th annual Biophysical Society Meeting: poster presentationJ.P Sumida, K Langsetmo, S.S. Lehrer. Conserved Asp137 destabilizes tropomyosin: CD, DSC and pressure perturbation measurements

50th annual Biophysical Society Meeting: poster presentationJ.P. Sumida, D. Hayes, K. Langsetmo, A. Hopping, S.S. Lehrer. Tropomyosin: Asp137→Leu Mutation Causes a Loss of Actin Binding.

51st annual Biophysical Society Meeting: poster presentation E.R. Wu, J.P. Sumida, S.S. Lehrer. Conserved Asp137 Locally Destabilizes Tropomyosin

Oral Presentations53rd annual Biophysical Society Meeting: poster presentation

J.P. Sumida, E.R. Wu, S.S. Lehrer. Conserved Destabilization of Tropomyosin at Asp 137: Functional Significance of Flexibility.

2008 Myofilament Meeting, (award talk).John Sumida, Eleanor Wu, David Hayes, Knut Langsetmo, Sherwin Lehrer. Tropomyosin: native instability and functional flexibility