Two-dimensional protein crystals. Annexin A5. |
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Geometric and Algebraic Order. 2D crystals are periodic arrangement of motiffs (atoms, molecules...) in two dimensions.
In the case of a three-dimensional crystal, the distances between the
motiffs are fixed (lattice constants). The order in 3D crystals is said
to be geometric.
In the case of a two-dimensional crystal, this is not the case: if one takes an arbitrary motiff as a
reference and measures the distance between it and any other motiff in
the crystal, one will find that that distance deviates from the
multiple of the lattice constant by an amount that increases
logarithmically with the distance. The
order in 2D crystals is said to be algebraic.
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This,
in turn, means that
diffraction peaks observed from 2D crystals are not delta-functions,
but power-laws. FWHM of the peak is a function of temperature and
elastic modulus of the crystal.
It also
follows that there is a limit
on resolution that can be obtained from diffraction studies 2D
crystals. It is, once again, related to the elastic properties of
the crystal. In practice, however, the resolution seems to be limited by
other factors.
Some of these issues are discussed in the work of Lenne et al. on the
grazing-angle X-ray diffraction studies from protein 2D crystals, Biophys. J. 79, 496-500 (2000),
and other publications by the same author.
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Main driving force behind gorowing and studying protein 2D crystlas is
their application in structural biology. They are used for
structure determination of soluble and transmembrane proteins by
electron crystallography.
A common procedure for growing 2D crystals of soluble proteins is the so-called
lipid monolayer method due to Kornberg et al.
In the case of membrane-binding proteins, the structure of the biologically
relevant, membrane-bound form, can be determined.
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Convenient
model systems for studying self-organisation, complex systems, and other soft condensed matter
physics phenomena;
Some may have biological role. In particular, ordered arrangements of
Annexin A5 are thought to assist in cell membrane repair: Bouter et al.
2010, Nature Communicaitons 2, 270.
Possbile applications in
biotechnology are also being investigated.
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Interesting link (web page of Jaap
Brink): List of proteins crystallised in two dimensions on lipid monolayers.
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Archetype of a family of soluble proteins expressed in many cell types in eucaryotes
that share structural homology (the so-called "annexin fold") and the
ability to bind lipids in a Ca2+ - dependent
manner.
Annexins participate in processes involving membrane fusion and trafficking, as
well as inhibition of blood coagulation.
It is now clear that the function of annexin A5 is in facilitating membrane repair (Bouter et al. 2010 Nature Communications 2, 270).
The ability of annexin A5 to bind to phospholipids has been used to grow 2D
crystals of this protein on lipid monolayers with the goal of
elucidating the structure of the membrane-bound form of the protein by
electron crystallography (see the work by Brisson et al. on this
subject).
The structure of the soluble form was solved by X-ray crystallography
(Huber et al., Lewit-Bentley et al).
Several forms of 2D crystals of annexin A5 exist. Initially two were known.
They differ with respect to their symmetry - p6 in one case and p3 in
the other. See the work of Dr. Frank Oling (Oling et al. 2001, J. Struct. Biol.13355-63) on this subject.
Some recent reviews on the subject: Gerke and Moss 2002 Physiological Reviews 82, 331-371 2002;
Moss and Morgan 2004, Genome Biology 5:Art. No. 219.
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1. Ca2+-dependent adsorption of the monomeric, soluble protein. 2. Fast
trimerisation step. 3. Nucleation and growth of p6 crystalline domains. 4. Solid-solid phase transition between the two crystal forms, p6 and p3.
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Annexin A5 2D crystals - p6
form.
B: 250x125 nm. Unit cell (blue lozenge): a=b=19.8 nm, Gamma: 1200.
C(36x36 nm), D(35x35 nm): Average 2D projection (calculated from EM
data) and average topography
(calculated from AFM data) maps, respectively. Here and on the right,
annexin V trimer is encircled in green. Numbers (pink) refer to
individual domains within the annexin A5
monomer. Numbering after Huber et al.
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Annexin A5 2D crystals - p3
form.
B (minimal force),
C(increased force) - 116 and 145 nm, respectively. Inset in B: average
topography map calculated by single particle averaging. D: Average topography
map of the p3 from calculated by single particle averaging methods.
3-fold symmetry imposed. 25 x 25 nm. E: 2D projection map of the p3
from calculated from EM data. Scale bar: 5 nm. Trimer-trimer
connections encircled in turquoise. |
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The Image on the left shows a small area of the p3
form, which appeared spontaneously within the p6 phase. A disordered
region (black asterisk) is seen to accompany its appearance. A vacancy
(missing annexin V trimer) is encircled in turquoise. |
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The image on the right shows defects - stacking
faults - which were found in the 2D crystals of the p6 crystal form.
The two examples highlighted with different colors exhibit a somewhat
different aspect. The omages in A is 546 x 458 nm, and the insets are
enlarged 2.2x. |
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The work shown here constitutes a part of my Doctoral thesis Atomic Force Microscopy of Biological Macromolecules and Their Assemblies. Advisor: Prof. A. Brisson, performed at the Department of Biophysical Chemistry, University of Groningen, the Netherlands.
It
would not be possible without both the previous and the con-current
structural characterization of the annexin A5 2D crystal forms by
electron crystallography the solution of its structure by X-ray
crystallography. Corresponding references and detailed
discssions can be found in the Thesis itself and inthe
relevant publications. Full list of Acknowledgments appears in the
Thesis. |
Background:
contact mode AFM Image of the p3 from of Annexin A5 2D crystals.
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