Professor of Physics
B.S., Oregon State University;
Ph.D., University of Oregon
Member of: Institute of Molecular Biology
Office: Willamette Hall Room 377
Lab: Willamette Hall Room 355
Our group uses an interdisciplinary approach in applying physical techniques to the study of biological molecules, especially the structure, function, and interaction of enzymes and fluorescent proteins. The primary technique we use is x-ray crystallography, but occasionally we do computer modeling of enzyme active sites and other properties of proteins. In the laboratory, chemists and biologists collaborate with physicists to achieve a broader intellectual basis for the research.
Green Fluorescent Protein
The newest and most exciting project in the laboratory involves a protein that spontaneously rearranges itself to become fluorescent, absorbing blue light (or UV) and re-emitting green light, hence the name Green Fluorescent Protein (GFP). GFP was discovered in the Pacific Northwest jellyfish Aequorea victoria and has become enormously popular as a visible tag for proteins of interest or as a marker for gene expression. It is nontoxic and has been expressed in essentially all types of organisms ranging from bacteria to mice. No host factors are required for the transformation to a fluorescent protein. If GFP is linked to a protein of interest, the cellular location of that protein in the living cell is revealed by a glance in the fluorescence microscope. We determined the structure of the protein in 1996 and have since embarked on a large project to generate a variety of biosensors by taking advantage of the fact that most forms of the protein actually have two absorption maxima that are sensitive to changes in the protein structure. Using genetic engineering techniques we have successfully constructed visual pH indicators, halide (chloride) concentration indicators and redox potential sensors. The color of the protein can also be modified by changing the environment or internal structure of the chromophore, which is derived from the primary sequence Ser(Thr)65-Tyr66-Gly67. We reported a yellow mutant in 1996 based on substitution of Thr203 with Tyr, but subsequently a Russian group has discovered related proteins from coral that fluoresce yellow and red, enabling multicolor reporting of a variety of cellular processes. It is fascinating that these different fluorescent proteins are nevertheless based on the same Xaa-Tyr-Gly peptide, and suggests that additional autocatalytic chemistry is involve in maturation of the protein. Crystals of a red variant are now on hand and work is well under way to determine the structure.
Enzyme Structure-Function Relationships
For many years we have worked to determine detailed structure function relationships in citrate synthase, which is at the entry to the citric acid cycle and is found in every organism examined. Citrate synthase, in its rate-determining step, abstracts a proton from the methyl group of acetylCoenzyme A to form a carbon-carbon double bond. The side chain which accomplishes this task is Asp375 working in concert with His274 (sequence numbering of pig heart enzyme). This equilibrium for this seemingly simple reaction is disfavored in solution by 12-15 orders of magnitude, and proposals for how an enzyme can do this are extremely controversial. Several publication have resulted from our studies, but the answer remains elusive. Recently, we determined the crystal structure of an enzyme that catalyzes an essentially identical reaction (malate synthase) in order to compare their respective mechanisms. It was fascinating to discover that the underlying chemistry is essentially the same in the two enzymes, but all of the details with the exception of an aspartic acid acting as a base are different. Evidently, Nature has discovered only one solution to this fundamental problem in chemistry, but the machinery is almost totally different! These studies are ongoing.
In the last few years, we have defined the first structures of two new families of enzymes, glycerol kinase and serine carboxypeptidase and may continue studies in these areas in the future. However, they are now "back burner" projects in favor of other exciting developments. Both enzymes are members of newly discovered superfamilies that are very diverse. For example glycerol kinase, actin and the heat shock cognate chaperonin (HSC70) are all ATPases with the same basic fold that utilize conformational changes upon hydrolysis of ATP to drive otherwise unrelated and extremely diverse biological processes.
Henderson JN, Ai HW, Campbell RE, Remington SJ. (2007) Structural basis for reversible photobleaching of a green fluorescent protein homologue. Proc Natl Acad Sci U S A. 104(16):6672-7.
Remington S.J. (2006) Fluorescent proteins: maturation, photochemistry and photophysics. Curr Opin Struct Biol, 16 (6): 714-21,
Remington S.J., R.M. Wachter, D.K. Yarbrough, B. Branchaud, D.C. Anderson, K. Kallio, and K.A. Lukyanov. (2005) zFP538, a yellow-fluorescent protein from Zoanthus, contains a novel three-ring chromophore. Biochem 44:202-12.
Dooley C.M., T.M. Dore, G.T. Hanson, W.C. Jackson, S.J. Remington, and R.Y. Tsien (2004) Imaging dynamic redox changes in mammalian cells with green fluorescent protein indicators. J Biol Chem Feb 25 [Epub ahead of print]
Rossignol R., R. Gilkerson, R. Aggeler, K. Yamagata, S.J. Remington, and R.A. Capaldi (2004) Energy substrate modulates mitochondrial structure and oxidative capacity in cancer cells. Cancer Res 64:985-93.
Hanson G.T., R. Aggeler, D. Oglesbee, M. Cannon, R.A. Capaldi, R.Y. Tsien, and S.J. Remington (2004) Investigating Mitochondrial Redox Potential with Redox-sensitive Green Fluorescent Protein Indicators. J Biol Chem 279:13044-53.
Anstrom D.M., K. Kallio, and S.J. Remington (2003) Structure of the Escherichia coli malate synthase G:pyruvate:acetyl-coenzyme A abortive ternary complex at 1.95 A resolution. Protein Sci 12:1822-32.
McAnaney T.B., E.S. Park, G.T. Hanson, S.J. Remington, and S.G. Boxer (2002) Green fluorescent protein variants as ratiometric dual emission pH sensors. 2. Excited-state dynamics. Biochem 41:15489-94.
Hanson G.T., T.B. McAnaney, E.S. Park, M.E. Rendell, D.K. Yarbrough, S. Chu, L. Xi, S.G. Boxer, M.H. Montrose, and S.J. Remington (2002) Green fluorescent protein variants as ratiometric dual emission pH sensors. 1. Structural characterization and preliminary application. Biochem 41:15477-88.
Capaldi, R.A., R. Aggeler, R. Gilkerson, G. Hanson, M. Knowles, A. Marcus, D. Margineantu, M. Marusich, J. Murray, D. Oglesbee, S.J. Remington, and R. Rossignol (2002) A replicating module as the unit of mitochondrial structure and functioning. Biochim Biophys Acta 1555:192-5.
De Giorgi F., L. Lartigue, M.K. Bauer, A. Schubert, S. Grimm, G.T. Hanson, S.J. Remington, R.J. Youle, and F. Ichas (2002) The permeability transition pore signals apoptosis by directing Bax translocation and multimerization. FASEB J 16:607-9.
Remington, S.J. (2002) Negotiating the speed bumps to fluorescence. Nat Biotechnol 20:28-9.
Yarbrough, D., R.M. Wachter, K. Kallio, M.V. Matz, and S.J. Remington (2001) Refined crystal structure of DsRed, a red fluorescent protein from coral, at 2.0-A resolution. PNAS 98:462-7.
Wachter, R. M., D. Yarbrough, K. Kallio, and S.J. Remington (2000) Crystallographic and energetic analysis of binding of selected anions to the yellow variants of green fluorescent protein. J Mol Biol 98:462-7.
Howard, B. R., J.A. Endrizzi, and S.J. Remington (2000) Crystal structures of Escherichia coli malate synthase G complexed with magnesium and glyoxylate at 2.0 A resolution: Mechanistic implications. Biochemistry 39:3156-68.
Jayaraman, S., P. Haggie, R.M. Wachter, S.J. Remington, and A.S. Verkman (2000) Mechanism and cellular applications of a green fluorescent protein-based halide sensor. J Biol Chem 275:6047-50.
Wachter, R.M. and S.J. Remington (1999) Sensitivity of the yellow variant of green fluorescent protein to halides and nitrate. Current Biology 9:R628-9.
Elsliger, M.A., R.M. Wachter, G.T. Hanson, K. Kallio, and S.J. Remington (1999) Structural and spectral response of green fluorescent protein variants to changes in pH. Biochemistry 38:5296-301.
Bystrom, C.E., D.W. Pettigrew, B.P. Branchaud, P. O'Brien, and S.J. Remington (1998) Crystal structures of Escherichia coli glycerol kinase mutant S58W in complex with non-hydrolyzable ATP analogs reveal a putative active conformation of the enzyme as a result of domain motion. Biochemistry 38:3508-18.
Usher, K.C., I.F. De la Cruz, F.W. Dahlquist, R.V. Swanson, M.I. Simon, and S.J. Remington (1998) Crystal structures of CheY from Thermotoga maritima do not support conventional explanations for the structural basis of enhanced thermostability. Protein Science 7:403-12.
Ormo, M., C.E. Bystrom, and S.J. Remington (1998) Crystal structure of a complex of Escherichia coli glycerol kinase and an allosteric effector fructose-1,6-biphosphate. Biochemistry 37:16565-72.
Usher, C., L.C. Blaszczak, G.S. Weston, B.K. Shoichet, and S.J. Remington (1998) Three-dimensional structure of AmpC beta lactamase from Escherichia coli bound to a transition-state analogue: possible implications for the oxyanion hypothesis and for inhibitor design. Biochemistry 37:16082-92.