Isomorphous replacement (IR) is historically the most common approach to solving the
phase problem in
X-ray crystallography
X-ray crystallography is the experimental science of determining the atomic and molecular structure of a crystal, in which the crystalline structure causes a beam of incident X-rays to Diffraction, diffract in specific directions. By measuring th ...
studies of
protein
Proteins are large biomolecules and macromolecules that comprise one or more long chains of amino acid residue (biochemistry), residues. Proteins perform a vast array of functions within organisms, including Enzyme catalysis, catalysing metab ...
s. For protein crystals this method is conducted by soaking the crystal of a sample to be analyzed with a
heavy atom solution or
co-crystallization with the heavy atom. The addition of the heavy atom (or ion) to the structure should not affect the crystal formation or
unit cell dimensions in comparison to its native form, hence, they should be
isomorphic
In mathematics, an isomorphism is a structure-preserving mapping or morphism between two structures of the same type that can be reversed by an inverse mapping. Two mathematical structures are isomorphic if an isomorphism exists between the ...
.
Data sets from the native and heavy-atom derivative of the sample are first collected. Then the interpretation of the
Patterson difference map reveals the heavy atom's location in the unit cell. This allows both the
amplitude and the
phase of the heavy-atom contribution to be determined. Since the
structure factor of the heavy atom derivative (F
ph) of the crystal is the vector sum of the lone heavy atom (F
h) and the native crystal (F
p) then the phase of the native F
p and F
ph vectors can be solved geometrically.
:
The most common form is multiple isomorphous replacement (MIR), which uses at least two isomorphous derivatives. Single isomorphous replacement is possible, but gives an ambiguous result with two possible phases;
density modification is required to resolve the ambiguity. There are also forms that also take into account the
anomalous X-ray scattering of the soaked heavy atoms, called MIRAS and SIRAS respectively.
Development
Single isomorphous replacement (SIR)
Early demonstrations of isomorphous replacement in crystallography come from
James M. Cork,
, and others. An early demonstration of isomorphous replacement in crystallography came in 1927 with a paper reporting the x-ray crystal structures of a series of
alum
An alum () is a type of chemical compound, usually a hydrated double salt, double sulfate salt (chemistry), salt of aluminium with the general chemical formula, formula , such that is a valence (chemistry), monovalent cation such as potassium ...
compounds from Cork.
The
alum
An alum () is a type of chemical compound, usually a hydrated double salt, double sulfate salt (chemistry), salt of aluminium with the general chemical formula, formula , such that is a valence (chemistry), monovalent cation such as potassium ...
compounds studied had the general formula A
.B
.(SO
4)
2.12H
2O, where A was a monovalent metallic ion (
NH4+,
K+,
Rb+,
Cs+, or
Tl+), B was a trivalent metallic ion (
Al3+,
Cr3+, or
Fe3+) and S was usually sulfur, but could also be
selenium
Selenium is a chemical element; it has symbol (chemistry), symbol Se and atomic number 34. It has various physical appearances, including a brick-red powder, a vitreous black solid, and a grey metallic-looking form. It seldom occurs in this elem ...
or
tellurium. Because the alum crystals were largely isomorphous when the heavy atoms were changed out, they could be phased by isomorphous replacement. Fourier analysis was used to find the heavy atom positions.
The first demonstration of isomorphous replacement in protein crystallography was in 1954 with a paper from
David W. Green,
Vernon Ingram, and
Max Perutz.
Multiple isomorphous replacement (MIR)
Examples
Some examples of heavy atoms used in protein MIR:
*
Hg2+ ions bind to
thiol groups.
*
Uranyl salts (
UO2 +
NO3) bind between
carboxyl groups in
Asp and
Glu
*
Lead
Lead () is a chemical element; it has Chemical symbol, symbol Pb (from Latin ) and atomic number 82. It is a Heavy metal (elements), heavy metal that is density, denser than most common materials. Lead is Mohs scale, soft and Ductility, malleabl ...
binds to Cys residues.
*
PtCl42− (
ion) bind to
His
See also
Anomalous diffraction
*
Multi-wavelength anomalous diffraction (MAD)
*
Single-wavelength anomalous diffraction (SAD)
Other
*
Patterson map
References
Further reading
*
*
External links
phase determination— a tutorial with illustrations and references.
Computer programs
SOLVE(now merged into PHENIX) – Terwilliger, T.C. and J. Berendzen. (1999) "Automated MAD and MIR structure solution". Acta Crystallographica D55, 849-861.
Tutorials and examples
{{Crystallography
X-ray crystallography