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by: Isidro Kuhlman


Isidro Kuhlman
OK State
GPA 3.83


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Class Notes
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This 0 page Class Notes was uploaded by Isidro Kuhlman on Sunday November 1, 2015. The Class Notes belongs to BIOC 6740 at Oklahoma State University taught by Staff in Fall. Since its upload, it has received 40 views. For similar materials see /class/232857/bioc-6740-oklahoma-state-university in Biochemistry at Oklahoma State University.

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Date Created: 11/01/15
Course Outline BIOC 6740 2008 Topics in Biological Spectroscopy I Review on Interaction of Radiation and Matter Properties of Electromagnetic Waves Frequency phase polarization Superposition of EM waves Plane circular and elliptically polarized radiation Coherence Interaction of Electromagnetic Radiation with Matter The dipole oscilator Scattering Re ection and Refraction Polarizability and refractive index Dispersion curves and birefringence Absorption of Light Energy of Radiation and Transitions The Transition Dipole Moment and the Probability of Absorption The Direction of the Transition Dipole Moment Polarized Absorption Spectra Einstein Relationships for Absorption and Emission of Radiation Boltzmann39s Distibution LambertBeer Law Lasers II UVVisible Absorption Spectroscopy Theory Rate of Electronic Transitions FrankCondom Principle Spectrum Shape Linewidth Natural and the uncertainty principle Effect of Intermolecular Interactions on Linewidth Effect of Intermolecular Interactions on the Energy of the Transition Solventinduced spectral shifts Transitions in the carbonyl and amide chromophores Interactions between chromophores Band splitting and Changes in Intensity Instrumental Aspects of UV Visible Absorption spectroscopy Single Beam Spectrophotometers SplitBeam Spectrophotometers Dual wavelength Spectrophotometer Diode Array Spectrophotometer Resolution accuracy and sensitivity of a spectrophotometric measurement Effect of stray light and Noise Comparison Different Spectrophotometers Resolution accuracy precision speed and sensitivity Analytical Methods used in UV Visible Absorption Spectroscopy Sample preparation and stability Selection of cuvette Temperature control effect on pH Single Wavelength determination of concentrations Multicomponent samples and the use of Multiple wavelengths Isosbestic Points UV absorption Spectra of Proteins and chromophores of biological importance Difference Spectra Derivative Spectroscopy III Fluorescence Spectroscopy 1 Principles Jablonsky Diagram Nonradiative Relaxation of Electronically Excited Molecules Vibrational Relaxation and Internal Conversion Deactivation 0f the thermallyequilibrated lowest excited singlet state radiative and nonradiative processes Fluorescence Singlettriplet Intersystem Crossing Phosphorescence Delayed Fluorescence Rates of Different Processes of Deactivation Stokes Shift Fluorescence Lifetime Quantum Yields 2 Steady State Fluorescence Spectroscopy The steadystate spectro uorometer Excitation and emission spectra Acquisition and Corrections Steady State Fluorescence Intensity Effects of quantum yield absorbance intensity of exciting radiation and slit widths Relationship between uorophore concentration and fluorescence intensity Ideal Relationship Inner Filter Effect Experimental Considerations Concentration of Fluorophores and Chromophores Scattering Rayleigh and Raman Selecting the Excitation Wavelength Sensitivity of uorescence comparison with UV absorption techniques 3 Fluorescence in Proteins Most common Fluorophores Phe Tyr and Trp Applications to study environment of uorophores Position of Maximum and Quantum Yields Effect of pH denaturants and Environment 4 Fluorescence Quenching Collisional Quenching The StemVolmer Equation Steady State and Lifetime Studies of Solute Quenching Static Quenching Applications of quenching studies Study of dynamic and structural properties of macromolecules ligandbinding and lipidprotein interaction 5 Fluorescence Polarization De nition of Anisotropy and Polarization Ratio Photoselection The emission transition dipole moment Anisotropy of Florescence Depolarization by Orientation Dependent Absorption Depolarization by noncolinearity between absorption and emission dipoles Depolarization by Molecular Motions Time Dependent Anisotropy of Fluorescence Steady State Anisotropy of Fluorescence Perrin s equation Macromolecular size and shape Study of chromophore dynamics in membranes and proteins Studies on LigandProtein and proteinprotein association Experimental Considerations The Gfactor 6 Resonance Energy Transfer Forster Theory The overlap integral The K factor Forster s Distance Applications Measurements of intermolecular distances proteinprotein interaction membraneprotein interactions Ligandbinding topology of membrane proteins lipidexchange between membranes and between proteins and membranes 7 Time resolved Fluorescence Spectroscopy Why measuring a uorescence lifetime Theory of FrequencyDomain uorometry Data Analysis Principle of frequencydomain measurements Instrumentation Light modulator crosscorrelation Frequency synthesizers and radio frequency amplifiers Applications of time resolved measurements in FRET and in quenching studies The measurement of Lifetimesdistributions using frequencydomain uorometry VI Some Biochemical Applications of UV Absorption Spectroscopy Protein folding Stability and kinetics of folding LigandBinding Experimental and theoretical approaches for single site and multiple equilibria DNA melting Chromophore Environment


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