The work proposed will develop UV resonance Raman spectroscopy (UVRS) as an incisive technique to examine protein structure and function. We will apply this technique for fundamental studies of amide, peptide and protein excited states and will search for delocalized electronic transitions similar to our recently discovered peptide charge transfer transitions. We will use a photochemical isomerization switching mechanism to determine the amino acid side chain dependence of the ground state trans yields cis peptide activation barriers. We will develop and utilize static and kinetic nsec-msec UVRS methods to examine the unfolding intermediates of proteins such as myoglobin (Mb) and apoM. We will, characterize the dynamical connectivities between adjacent peptide bonds by selectively initiating trans yields cis isomerization in a thioamide substituted peptide, and then measure the conformational changes at adjacent peptide bonds. We will improve UV Raman instrumentation, to increase its sensitivity.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM030741-18
Application #
6180322
Study Section
Molecular and Cellular Biophysics Study Section (BBCA)
Program Officer
Wehrle, Janna P
Project Start
1982-06-01
Project End
2002-06-30
Budget Start
2000-07-01
Budget End
2001-06-30
Support Year
18
Fiscal Year
2000
Total Cost
$270,971
Indirect Cost
Name
University of Pittsburgh
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
053785812
City
Pittsburgh
State
PA
Country
United States
Zip Code
15213
Lanoul, Anatoli; Coleman, Thomas; Asher, Sanford A (2002) UV resonance raman spectroscopic detection of nitrate and nitrite in wastewater treatment processes. Anal Chem 74:1458-61
Asher, S A; Ianoul, A; Mix, G et al. (2001) Dihedral psi angle dependence of the amide III vibration: a uniquely sensitive UV resonance Raman secondary structural probe. J Am Chem Soc 123:11775-81
Lednev, I K; Karnoup, A S; Sparrow, M C et al. (2001) Transient UV Raman spectroscopy finds no crossing barrier between the peptide alpha-helix and fully random coil conformation. J Am Chem Soc 123:2388-92
Boyden, M N; Asher, S A (2001) UV Raman studies of peptide conformation demonstrate that betanova does not cooperatively unfold. Biochemistry 40:13723-7
Ianoul, A; Boyden, M N; Asher, S A (2001) Dependence of the peptide amide III vibration on the phi dihedral angle. J Am Chem Soc 123:7433-4
Holtz, J S; Lednev, I K; Asher, S A (2000) UV resonance Raman study of angiotensin II conformation in nonaqueous environments: lipid micelles and acetonitrile. Biopolymers 57:55-63
Holtz, J S; Holtz, J H; Chi, Z et al. (1999) Ultraviolet Raman examination of the environmental dependence of bombolitin I and bombolitin III secondary structure. Biophys J 76:3227-34
Chi, Z; Asher, S A (1999) Ultraviolet resonance Raman examination of horse apomyoglobin acid unfolding intermediates. Biochemistry 38:8196-203
Chi, Z; Asher, S A (1998) UV resonance Raman determination of protein acid denaturation: selective unfolding of helical segments of horse myoglobin. Biochemistry 37:2865-72
Chi, Z; Chen, X G; Holtz, J S et al. (1998) UV resonance Raman-selective amide vibrational enhancement: quantitative methodology for determining protein secondary structure. Biochemistry 37:2854-64

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