Abstract
The chemical nature of the enzyme-substrate intermediates studied by resonance Raman (RR) spectroscopy is defined. The information available on these intermediates from RR spectroscopy is compared to that obtained from absorption and fluorescence techniques. Two major classes of substrates useful for RR studies are discussed. The first class involves the use of substrates which are chromophoric by virtue of the fact that they have acyl groups, e.g. cinnamoyl, based on delocalised π-systems. Their use to probe the effect on the substrate’s C=0 group of going to active-pH in serine proteases, and to probe the strong electron polarisation forces available in cysteine proteases, is detailed. The second class of substrate involves thionoesters RC(=S)OCH, which form chromophoric dithioester intermediates, RC(=S)S-enzyme, with cysteine proteases, HS-enzyme. Because the dithioester RR spectrum is sensitive to conformational changes in the R-C(=S)-S-C bonds it is used to characterise conformational activation in the scissile linkages which occur for substrates specific for the cysteine protease papain. In addition RR spectra of these dithio-intermediates have been obtained under cryoenzymological conditions at 77°K and offer the opportunity to characterise such dynamical events as the thermal activation of the substrate in the active-site.
Keywords
- Ground Electronic State
- Resonance Raman
- Resonance Raman Spectrum
- Tetrahedral Intermediate
- Resonance Raman Spectroscopy
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.
Published as NRCC No. 29252
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Preview
Unable to display preview. Download preview PDF.
References
P. D. Boyer (Ed.) “The Enzymes” Vol 3 Academic Press, N.Y. (1971).
A. Fersht, “Enzyme Structure and Mechanism”, W.H. Freeman, N.Y. (1985).
P. R. Carey, “Biochemical Applications of Raman and Resonance Raman Spectroscopies”, Academic Press, N.Y. (1982).
B. A. E. MacClement, R. G. Carriere, D. J. Phelps and P. R. Carey, Biochemistry, 20, 3438–3447 (1981).
P. R. Carey, “Resonance Raman Labels and Enzyme-Substrate Reactions”, in “Biological Applications of Raman Spectrometry”, Vol. 2, (T.G. Spiro, Ed) J. Wiley, N.Y. (1987).
I. D. Campbell and R. A. Dweck, “Biological Spectroscopy”, Chapter 5, Benjamin-Cummings, Menlo Park, Calif. (1984).
S. A. Bernhard and S.-J. Lau, “Cold Spring Harbor Symp. Quant. Bol. 36, 75–83 (1972).
A. Warshel and S. T. Russell, Q. Rev. Biophys. 17, 283–422 (1984).
P. R. Carey and A. C. Storer, Ann. Rev. Biophys. Bioeng., 13, 25–49 (1984).
J. B. Henes, J.A. Mattis and J. S. Fruton, Proc. Natl. Acad. Sci. USA, 76, 1131–1134 (1979).
P. R7-Carey, R. G. Carriere, D. J. Phelps and H. Schneider, Biochemistry, 17, 1081–1087 (1978).
A. C. Storer, W. F. Murphy and P. R. Carey, J. Biol. Chem. 254, 3163–3165 (1979).
P. R. Carey and A. C. Storer, Pure Appl. Chem., 57, 225–246 (1985).
A. C. Storer and P. R. Carey, Biochemistry, 24, 6808–6818 (1985).
R. H. Angus, P. R. Carey, H. Lee and A. C. Storer, Biochemistry, 25-, 3304–3310 (1986).
C. P. Huber, Acta Cryst. C43, 902–904 (1987).
K. I. Varughese, C. P. Huber, A. C. Storer and P.R. Carey, ms. in preparation.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 1989 Springer Science+Business Media New York
About this chapter
Cite this chapter
Carey, P.R. (1989). Using Resonance Raman Spectroscopy to Study the Structure and Dynamics of Enzyme-Bound Substrates. In: Cooper, A., Houben, J.L., Chien, L.C. (eds) The Enzyme Catalysis Process. Progress in Mathematics. Springer, Boston, MA. https://doi.org/10.1007/978-1-4757-1607-8_13
Download citation
DOI: https://doi.org/10.1007/978-1-4757-1607-8_13
Publisher Name: Springer, Boston, MA
Print ISBN: 978-1-4757-1609-2
Online ISBN: 978-1-4757-1607-8
eBook Packages: Springer Book Archive