Abstract
Elucidating proteins function at a level that allows for intelligent design and manipulation is essential in realization of their potential role in biomedical and industrial applications. It has become increasingly apparent though, that probing structures and functionalities under equilibrium conditions is not sufficient. Rather, many aspects of protein behavior and reactivity are rooted in protein dynamics. Thus, there is a growing effort to probe intermediate structures that occur transiently during the course of a proteins function in particular linked to the binding or release of a ligand or substrate. However, studies following the sequence of conformational changes triggered by the binding of sub-strate/ligand and the concomitant change in functional properties are inherently difficult because often the diffusion times are of the order of conformational relaxation times. This chapter describes methodologies for generating resonance Raman spectra from transient forms of hemoglobin under conditions that allow for the systematic exploration of conformational relaxation and functionality. Special consideration is given to Raman compatible protocols based on sol-gel encapsulation that allow for the preparation, trapping and temporal tuning of nonequilibrium population generated from either the addition or the removal of ligands/substrates.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Spiro T. G. (1978) Resonance Raman spectra of hemoproteins. Methods Enzymol 54, 233–249.
Spiro T. G. (1985) Resonance Raman spectroscopy as a probe of heme protein structure and dynamics. Adv. Protein Chem. 37, 111–159.
Spiro T. G., Smulevich G., and Su C. (1990) Probing protein structure and dynamics with resonance Raman spectroscopy: cytochrome c peroxidase and hemoglobin. Biochemistry 29, 4497–4508.
Spiro T. G. and Czernuszewicz R. S. (1995) Resonance Raman spectroscopy of metalloproteins. Methods Enzymol. 246, 416–460.
Friedman J. M. (1994) Time-resolved resonance Raman spectroscopy as probe of structure, dynamics, and reactivity in hemoglobin. Methods Enzymol. 232, 205–231.
Asher S. (1993) UV resonance Raman spectroscopy for analytical, physical and biophysical chemistry. Part 1. Anal. Chem. 65, 59A–66A.
Austin J., Jordan T., and Spiro T. (1993) Ultraviolet resonance Raman studies of proteins and related model compounds. In Biomolecular Spectroscopy Part A (Clark R. J. H. and Hester R. E., eds.), John Wiley and Sons, New York, pp. 55–127.
Kitagawa T. (1988) The heme protein structure and the iron histidine stretching mode. In Biological Application of Raman Spectroscopy, Vol. III (Spiro T. G., ed.), John Wiley & Sons, New York, pp. 97–131.
Rousseau D. L. and Friedman J. M. (1988) Transient and cryogenic studies of photodissociated hemoglobin and myoglobin. In Biological Applications of Raman Spectroscopy, Vol. III (Spiro T. G., ed.), John Wiley & Sons, New York, pp. 133–215.
Friedman J. M., Scott T. W., Stepnoski R. A., Ikeda-Saito M., and Yonetani T. (1983) The iron-proximal histidine linkage and protein control of oxygen binding in hemoglobin. A transient Raman study. J. Biol. Chem. 258, 10,564–10,572.
Friedman J. M. (1985) Structure, dynamics, and reactivity in hemoglobin. Science 228, 1273–1280.
Scott T. W. and Friedman J. M. (1984) Tertiary-structure relaxation in hemoglobin: a transient Raman study. J. Am. Chem. Soc. 106, 5677–5687.
Avnir D., Braun S., Lev O., and Ottolenghi M. (1994) Enzymes and other proteins entrapped in sol-gel materials. Chem. Mater. 6, 1605–1614.
Bettati S. and Mozzarelli A. (1997) T state hemoglobin binds oxygen noncooperatively with allosteric effects of protons, inositol hexaphosphate, and chloride. J. Biol. Chem. 272, 32,050–32,055.
Bruno S., Bonaccio M., Bettati S., Rivetti C., Viappiani C., Abbruzzetti S., and Mozzarelli A. (2001) High and low oxygen affinity conformations of T state hemoglobin. Protein Sci. 10, 2401–2407.
Dave B. C., Miller J. M., Dunn B., Valentine J. S., and Zink J. I. (1997) Encapsulation of proteins in bulk and thin film sol-gel matrices. J. Sol Gel Sci. Technol. 8, 629–634.
Das T. K., Khan I., Rousseau D. L., and Friedman J. M. (1999) Temperature dependent quaternary state relaxation in sol-gel encapsulated hemoglobin. Biospectroscopy 5, S64–S70.
Ellerby L. M., Nishida C. R., Nishida F., Yamanaka S. A., Dunn B., Valentine J. S., and Zink J. I. (1992) Encapsulation of proteins in transparent porous silicate glasses prepared by the sol-gel method. Science 255, 1113–1115.
Juszczak L. J. and Friedman J. M. (1999) UV resonance Raman spectra of ligand binding intermediates of sol-gel encapsulated hemoglobin. J. Biol. Chem. 274, 30,357–30,360.
Khan I., Shannon C. F., Dantsker D., Friedman A. J., Perez-Gonzalezde-Apodaca J., and Friedman J. M. (2000) Sol-gel trapping of functional intermediates of hemoglobin: geminate and bimolecular recombination studies. Biochemistry 39, 16,099–16,109.
Samuni U., Dantsker D., Khan I., Friedman A. J., Peterson E., and Friedman J. M. (2002) Spectroscopically and kinetically distinct conformational popula 300 tions of sol-gel encapsulated carbonmonoxy myoglobin: a comparison with hemoglobin. J. Biol. Chem. 25, 25.
Shibayama N. and Saigo S. (1995) Fixation of the quaternary structures of human adult haemoglobin by encapsulation in transparent porous silica gels. J. Mol. Biol. 251, 203–209.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2005 Humana Press Inc., Totowa, NJ
About this protocol
Cite this protocol
Samuni, U., Friedman, J.M. (2005). Proteins in Motion. In: Ulrich Nienhaus, G. (eds) Protein-Ligand Interactions. Methods in Molecular Biology, vol 305. Humana, Totowa, NJ. https://doi.org/10.1385/1-59259-912-5:287
Download citation
DOI: https://doi.org/10.1385/1-59259-912-5:287
Publisher Name: Humana, Totowa, NJ
Print ISBN: 978-1-58829-372-5
Online ISBN: 978-1-59259-912-7
eBook Packages: Springer Protocols