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Chapter 3
New Insight into Solution Structure and Dynamics of Proteins, Nucleic Acids, and Viruses from Raman Optical Activity L. D. Barron , E. W. Blanch , A. F. Bell , C. D. Syme , 1
1
1
1
L. Hecht , and L. A. D a y 2 1
Department of Chemistry, University of Glasgow, Glasgow G12 8QQ, United Kingdom Public Health Research Institute, 455 First Avenue, New York, N Y 10016 1
2
Raman optical activity measures vibrational optical activity by means of a small difference in the intensity of Raman scattering from chiral molecules in right and left circularly polarized incident light. The sensitivity of R O A to chirality makes it an incisive probe of biomolecular structure and dynamics in aqueous solution. This article reviews the basic theory and instrumentation of R O A , and describes recent results which illustrate how R O A provides new insight into current biomedical problems including protein misfolding and disease, and virus structure at the molecular level.
Introduction Studies of the structure and dynamics of biomolecules in aqueous solution remains at the forefront of biomedical science. The potential value of vibrational spectroscopy, both infrared and Raman, has been greatly enhanced in this area
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© 2002 American Chemical Society
In Chirality: Physical Chemistry; Hicks, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 2002.
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by the addition of the new dimension of optical activity (1-3), which confers an exquisite sensitivity to the absolute stereochemistry and conformation of chiral molecules. The word 'chiral', meaning handed, was first introduced into science by Lord Kelvin (4), Professor of Natural Philosophy in the University of Glasgow. Phenomena which are sensitive to molecular chirality include optical rotation, and ultraviolet circular dichroism (UVCD) where left and right circularly polarized U V light is absorbed slightly differently. Such 'chiroptical' techniques have a special sensitivity to the three dimensional structure of a chiral molecule and are widely used in the conformational analysis of biomolecules. The importance of the newer vibrational optical acivity methods is that they are sensitive to chirality associated with all 3N-6 fundamental vibrational transitions, where Ν is the number of atoms, and therefore have the potential to provide much more stereochemical information than U V C D which measures optical activity associated with the far fewer accessible electronic transitions. Vibrational optical activity in typical chiral molecules in the liquid phase was.first observed by Barron et al in 1973 (5) using a Raman optical activity technique in which, as depicted in Figure 1, a small difference in the intensity of Raman scattering is measured using right- and left-circularly polarized incident light. Until recently, however, lack of sensitivity restricted R O A to favourable samples such as small chiral organic molecules. But major advances in instrumentation have, in recent years, rendered biomolecules in aqueous solution accessible to R O A studies (6). Conventional Raman spectroscopy has a number of favourable characteristics which have led to many applications in biochemistry. In particular, the complete vibrational spectrum from -100 to 4000 cm" is accessible on one simple instrument, and both H2O and D 0 are excellent solvents for Raman studies. Since R O A is sensitive to chirality, it is able to build on these advantages by adding to Raman spectroscopy an extra sensitivity to the three-dimensional structure, which opens a new window on biomolecular problems (7). !
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Theory of R O A
The fundamental scattering mechanism responsible for R O A was discovered by Atkins and Barron in 1969 (8), who showed that interference between waves scattered via the polarizability and optical activity tensors of a chiral molecule yields a dependence of the scattered intensity on the degree of circular
In Chirality: Physical Chemistry; Hicks, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 2002.
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R
I -I *O R
L
incident beam with angular frequency ω
Raman component of scattered beam with angular frequency ω -ω ν
Figure L The basic ROA experiment measures a small difference in the intensity of Raman scattering from chiral molecules in right- and left-circularly polarized incident light.
In Chirality: Physical Chemistry; Hicks, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 2002.
37 polarization of the incident light and to a circular component in the scattered light. Barron and Buckingham (9) subsequently developed a more complete version of the theory and introduced the following definition of the dimensionless circular intensity difference (CID) 4 = (/*-/ )/(/*+/ ) L
L
as an appropriate experimental quantity, where I and I are the scattered intensities in right- and left-circularly polarized incident light. In terms of the electric dipole-electric dipole molecular polarizability tensor α and the
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R
L
αβ
electric dipole-magnetic dipole and electric dipole-electric quadrupole optical activity tensors & and Α (1,2,9), the CIDs for forward (0°) and backward α β
αβγ
(180°) scattering from an isotropic sample are as follows: 4(0°)=4[45aG'+j8(G ) -/?(a) j/c[45a +7j3(a) ] /
2
2
2
4(180°)= 24|j3(G') + \β{Α) ]/^45α 2
2
2
+ Ίβ(α) \
2
2
where the isotropic invariants are defined as a
3 aa>
=
G'
a
~\ 'aa G
and the anisotropic invariants as β(α)
2
= 1 {3α α αβ
αβ
-~α α αα
)
ββ
β(Α)
2
β(θ')
2
= \{3α Ο' αβ
-α Ο'
αβ
αα
ββ
\
-\(οα ε Α αβ
αγ8
γδβ
We are using a Cartesian tensor notation in which a repeated Greek suffix denotes summation over the three components, and Β is the third-rank unit αβγ
antisymmetric tensor. These results apply specifically to Rayleigh scattering. For Raman scattering the same basic C I D expressions apply but with the molecular property tensors replaced by corresponding vibrational Raman transition tensors. Using a bond polarizability theory of R O A for the case of a molecule composed entirely of idealized axially symmetric bonds, the relationships
In Chirality: Physical Chemistry; Hicks, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 2002.
38 β(β') =β{Α) 2
2
and a G ' = 0 are found (1). Within this model,
therefore,
isotropic scattering makes zero contribution to the R O A intensity which is generated exclusively by anisotropic scattering, and the forward and backward CIDs reduce to
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4(0°)=0,
4l80°)=32j3(G ) /4 +0(
