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domains of unknown function. We also show how minor changes in otherwisehighly conserved active sites can significantly affect functionality. There isa growing need for intelligent prediction-based strategies that can tap intoour enormous genomic and structural databases and help bridge the gap be-tween sequence and function.

1183-Pos Board B75Epileptic Seizures-Induced Structural Changes in Rat Spine Bone Tissues:FTIR Microspectroscopic and Chemometric StudySebnem Garip1,2, Deniz Sahin3,4, Mete Severcan1,5, Feride Severcan1,6.1Middle East Technical University, Ankara, Turkey, 2Department ofBiochemistry, Middle East Technical University, Ankara, Turkey, 3KocaeliUniversity Faculty of Medicine, Kocaeli, Turkey, 4Department ofPhysiology, Kocaeli University Faculty of Medicine, Kocaeli, Turkey,5Department of Electrical and Electronics Engineering, Middle EastTechnical University, Ankara, Turkey, 6Department of Biological Sciences,Middle East Technical University, Ankara, Turkey.Epilepsy is a common serious neurodegenerative disease. Bone disorders due toanti-epileptic drug (AED) therapy in epileptic patients have been reported pre-viously. There is no study in the literature, investigating the independent effectof epileptic seizures on bone tissues. Thus, the side-effects of AEDs on bonetissues could not be differentiated from the effects of the epileptic seizures.The current study provides the first report on clarifying the effects of seizureson bones. The experiments performed on genetically epileptic and healthy rats,give the advantage of studying the effects of seizures alone. Cortical region ofspines were studied by FTIR microscopy to investigate the structural and com-positional changes in bones. Comparison of FTIR images belonged to the min-eral and protein parts of bone clearly showed the difference between healthyand epileptic bone tissues. Mineral content was found to be decreased in epilep-tic group compared to the healthy control. Although total carbonate content wasfound to be decreased, B-type carbonate content which substitutes for phos-phate groups in the mineral part of bone, was shown to be increased in epilepticgroup. The organic matrix of bone is mainly composed of collagen proteinswhose structure is stabilized by several intermolecular crosslinks. Collagencross-links ratio was found to be changed critically in epileptic group, indicat-ing an increase in immature crosslinks in the bones of that group. Crystallinityvalue indicating crystal size was found to be increased in epileptic group com-pared to the healthy control. Decreased mineral content and collagen crosslinksand increased crystal size and carbonate substitution, imply a severe damage onbone tissues. Moreover, the epileptic and control groups were separated fromeach other successfully by principle component analysis (PCA) based on theFTIRM data.

1184-Pos Board B76A Spectroscopic Survey of Substituted Indoles Reveals Effects of 1LB

Transition StabilizationXianwei Meng1, Trisheena Harricharran2, Laura J. Juszczak1.1Brooklyn College/The City University of New York, Brooklyn, NY, USA,2The Graduate Center/The City University of New York, New York, NY,USA.Although tryptophan is a natural probe of protein structure, interpretation ofits fluorescence emission spectrum is complicated by the presence of twoelectronic transitions, 1La and 1Lb. Theoretical calculations show thata point charge adjacent to either ring of the indole can shift the emissionmaximum. This study explores the effect of pyrrole and benzyl ring substitu-tions on the transitions’ energy via absorption and fluorescence spectroscopy,and lifetime measurements. The survey of indole derivatives shows thatmethyl substitutions on the pyrrole ring effect 1La and

1Lb energies in tandemwhile benzyl ring substitutions with electrophilic groups lift the 1La/

1Lb

degeneracy. For 5- and 6-hydroxyindole in cyclohexane, 1La and1Lb transi-

tions are resolved (5-hydroxyindole absor-bance, shown, solid line). This findingprovides for 1La origin assignment in theabsorption and excitation spectra for in-dole vapor. The 5-hydroxyindole excita-tion spectrum (dashed line) shows thatdespite a blue-shifted emission spectrum,both the 1La and

1Lb transitions contributeto emission. 10 0 ns fluorescence lifetimesfor 5-hydroxyindole are consistent with acharge acceptor-induced increase in thenonradiative rate.

Enzymes

1185-Pos Board B77Use Nanomechanical Sensor to Detect Cellulase Activities IncludingEnzymatic Decrystallization and Hydrolytic Cleavage on CelluloseWenjian Du, Liming Zhao, Chi Nguyen, Jun Xi.Drexel University, philadelphia, PA, USA.Cellulase is an interfacial enzyme that catalyzes the hydrolytic degradation ofcellulose at the interface between a liquid phase (enzyme) and a solid phase(cellulose substrate). Prior to the hydrolytic cleavage, cellulase utilizes an ac-tivity known as enzymatic decrystallization to break up the solid aggregateof cellulose molecules. The activity of enzymatic decrystallization has notbeen characterized and its mechanism has not been elucidated because veryfew existing experimental approaches are able to examine interfacial enzymaticactivity on solid substrates. Here, we report the development of a novel strategyfor the real-time detection of cellulase activities including enzymatic decrystal-lization and hydrolytic cleavage on cellulose with the use of a nanomechanicalsensor in a microcantilever. We present both kinetic and physical evidence tosupport the decrystallization as a kinetically viable step of cellulose hydrolysisby cellulase. To our knowledge, this is the first use of a nanomechanical sensorto study mechanistic enzymology and heterogeneous enzymatic catalysis thatinvolves a solid substrate. This nanomechanical sensor-based approach willhelp obtain a comprehensive understanding of cellulase actions on cellulose,which would be essential to the success of the development of new cellulaseswith enhanced efficiency for biofuels production.

1186-Pos Board B78Single Enzyme Studies Reveal the Existence of Discrete Functional Statesfor Monomeric Enzymes and How they are ‘‘Selected’’ upon AllostericRegulationNikos S. Hatzakis1, Li Wei1, Sune K. Jorgensen1, Andreas H. Kunding1,Pierre-Yves Bolinger1, Nicky Ehrlich1, Ivan Makarov1, Michael Skjot2,Allan Svendsen2, Per Hedegard3, Samuel M. Walsh1, Dimitrios Stamou1.1University of Copenhagen, Copenhagen, Denmark, 2Novozymes A/S,Department of Protein Biochemistry, Copenhagen, Denmark, 3Nano-ScienceCenter, Niels Bohr Institute, University of Copenhagen, Copenhagen,Denmark.Allosteric regulation of enzymatic activity forms the basis for controllinga plethora of vital cellular processes. While the mechanism underlying regula-tion of multimeric enzymes is generally well understood and proposed to pri-marily operate via conformational selection, the mechanism underlyingallosteric regulation of monomeric enzymes is poorly understood. Here wemonitored for the first time allosteric regulation of enzymatic activity at the sin-gle molecule level (1). We measured single stochastic catalytic turnovers ofa monomeric metabolic enzyme (Thermomyces lanuginosus Lipase) while ti-trating its proximity to a lipid membrane that acts as an allosteric effector.The single molecule measurements revealed the existence of discrete binaryfunctional states that could not be identified in macroscopic measurementsdue to ensemble averaging. The discrete functional states correlate with the en-zyme’s major conformational states and are redistributed in the presence of theregulatory effector. Thus, our data support allosteric regulation of monomericenzymes to operate via selection of preexisting functional states and not via in-duction of new ones.(1) Hatzakis, N. S.; Wei, L.; Jørgensen, S. K.; Kunding, A., H.; Bolinger, P-Y.;Ehrlich, N.; Makarov, I.; Skjot, M.; Svendsen, A.; Hedegard, P.; Stamou, D.(2012). Single Enzyme Studies Reveal the Existence of Discrete FunctionalStates for Monomeric Enzymes and How They Are ‘‘Selected’’ upon AllostericRegulation. J. Am. Chem. Soc.134 (22), 9296-9302.

1187-Pos Board B79Backbone 1H-13C-15N NMR Assignments and Ligand Binding Study ofOMP Synthase from Saccharomyces CerevisiaeMichael R. Hansen1, Eric W. Barr1, Richard Harris2, Kaj Frank Jensen3,Martin Willemoes3, Hong Cheng4, Mark Girvin2, Charles Grubmeyer1.1Temple University School of Medicine, Philadelphia, PA, USA, 2AlbertEinstein College of Medicine of Yeshiva University, Bronx, NY, USA,3University of Copenhagen, Copenhagen, Denmark, 4Fox Chase CancerCenter and Temple University School of Medicine, Philadelphia, PA, USA.Catalysis in OMP synthase (orotate phosphoribosyltransferase, EC 2.4.2.10) iscoupled to the unstructured-to-structured transition of a 10-residue peptideloop. OMP synthase from yeast is a small, homodimeric (49 kDa) and highlystable domain-swapped enzyme that catalyzes the formation of the UMP