About the Project:
Iron-overload diseases are usually genetic and are extremely
common in many parts of the world. In areas such as South East Asia
they constitute a major public health problem. In Australia, the gene
for hemochromatosis is carried by about 1% of the population. Iron
accumulates in patients and precipitates in various parts of the body
in the form of iron oxyhydroxide particles. Some of our research over
the past few years has contributed to the discovery that there are
three major mineral structures of iron oxyhydroxide that can be found
in iron-overloaded tissues.
Most living systems are able to store iron in the form of nanoscale
particles of iron(III) oxyhydroxide encapsulated by the protein
ferritin. Each molecule of ferritin consists of 24 subunits which are
assembled to form an approximately spherical cage-like structure of
external diameter 12 nm [1]. The internal cavity of the structure,
within which the iron(III) oxyhydroxide particle is stored, has a
diameter of approximately 8 nm. Thus the cavity limits the number of
iron atoms in each particle to a maximum of approximately 4500. The
protein shell surrounding each particle renders it water-soluble. Under
some circumstances, e.g. pathological conditions of iron-overload,
insoluble iron(III) oxyhydroxide particles are found to be deposited in
biological tissue [2, 3]. This form of iron deposit is often termed
hemosiderin. Hemosiderin may be a degradation product of ferritin or
may be formed independently. Hemosiderin is not associated with well
formed protein cavities and as such the particle size distribution may
be less restricted when compared with ferritin.
The structure of the iron(III) oxyhydroxide particle of most ferritins
is based on that of the mineral ferrihydrite (5Fe2O3.9H2O) with varying
amounts of phosphate incorporated [2]. In some cases, such as the
bacterioferritins from Azotobacter vinelandii and Pseudomonas
aeruginosa, the phosphate content is large so that the composition is
better described as a hydrated iron(III) oxyphosphate [4, 5]. The
degree of crystallinity of the particles within ferritins can also vary
from the well ordered structure of ferrihydrite through to a
non-crystalline state as determined by electron diffraction
measurements. The degree of crystallinity and particle size
distribution of the inorganic particles in a native ferritin depend on
the source of the ferritin and have also been observed to vary with
pathological condition [6].
The variations in structure of the mineral particles in hemosiderins
are even more marked than those in ferritins [2, 7, 8]. Three different
particle structures have been identified, namely (i) a structure based
on that of ferrihydrite (ii) a highly defect structure based on that of
the mineral goethite (a-FeOOH), and (iii) a non-crystalline iron(III)
structure. The particle size distributions in hemosiderins are often
difficult to measure because of aggregation. However, transmission
electron microscope observations clearly indicate that the
distributions are generally broader than those of ferritins. Like
ferritins, the structural and compositional characteristics of
hemosiderin vary with biological source and pathological condition [2,
7-9].
The methodology used to assign these solid state structures to the
hemosiderins was based on the biochemical isolation of the hemosiderins
from tissue samples followed by characterisation using Mössbauer
spectroscopy and electron diffraction. All three structures have been
observed to occur in human tissues under varying pathological
conditions. This chemical speciation of hemosiderin iron oxides is of
importance because the different structures are expected (i) to have
different degrees of toxicity, (ii) to have different reactivities with
chelating drugs, (iii) to reflect different mechanisms of iron
deposition. Subsequent to the discovery of the three different
structures, several workers have used Mössbauer spectroscopy of frozen
or freeze dried tissue specimens to elucidate the structures of the
iron oxide deposits present in situ. This has proved to be a powerful
technique, allowing characterisation of iron oxide structures in many
more tissue specimens without the need for biochemically isolating the
iron oxide deposits from the tissue. The three different iron oxide
structures can be differentiated with the use of Mössbauer spectroscopy
by virtue of their different temperature dependent magnetic behaviour.
Thus, Mössbauer spectroscopic measurements generally need to be made at
several different sample temperatures in order to characterise the
magnetic behaviour sufficiently for determination of the structures
present. A disadvantage of the Mössbauer spectroscopic technique is
that each spectrum requires at least 24 hrs (and usually more) for data
collection. Thus one sample may require 1 week or more for full
characterisation.
We have made the first ever use of AC-magnetic susceptibility
measurements to characterise the temperature dependent magnetic
behaviour of hemosiderins [12]. We show that this technique yields
greater detail about the temperature dependent magnetic behaviour of
hemosiderins and that the data can be collected on a time-scale one
order of magnitude smaller than that required for Mössbauer
spectroscopy. This opens the way for larger scale studies of the
structure of iron oxide deposits in iron loaded tissues. In addition, a
detailed knowledge of the magnetic properties of pathological tissue
iron oxide deposits is necessary for the interpretation of magnetic
resonance images of iron loaded tissues [10, 11].
This project has been carried out in collaboration with Prof K.V.Rao
and Dr J.L. Costa-Krämer at the Royal Institute of Technology,
Stockholm, Prof J. Webb and D.J. Macey, Murdoch University, Perth, and
Prof P. Pootrakul, Mahidol University, Bangkok.
References:
- P.M. Harrison and P. Arosio. The ferritins: molecular
properties, iron storage function and cellular regulation, Biochim.
Biophys. Acta. 1275 (1996) 161-203.
- T.G. St. Pierre, J. Webb and S. Mann (1989) in
Biomineralization: chemical and biochemical perspectives (S. Mann, J.
Webb, R.J.P. Williams, eds.), pp. 295-344, VCH, Weinheim.
- T. Iancu. Ferritin and hemosiderin in pathological tissues, Electron Microsc. Rev. 5 (1992) 209-229.
- S. Mann, J.V. Bannister and R.J.P. Williams. Structure
and composition of ferritin cores isolated from human spleen, limpet
(patella vulgata) hemolymph and bacterial (Pseudomonas aeruginosa)
cells, J. Mol. Biol. 188 (1986) 225-232.
- G. Watt, R.B. Frankel, G.C. Papaefthymiou, K.
Spartalian and E.I. Stiefel. Redox Properties and Mössbauer
spectroscopy of Azotobacter vinelandii bacterioferritin, Biochem. 25
(1986) 4330-4336.
- T.G. St. Pierre, K.C. Tran, J. Webb, D.J. Macey, B.R.
Heywood, N.H. Sparks, V.J. Wade, S. Mann and P. Pootrakul. Organ
specific crystalline structures of ferritin cores in
b-thalassaemia/haemoglobin E, Biol. Metals 4 (1991) 162-165
- D.P.E. Dickson, N.M.K. Reid, S. Mann, V.J. Wade, R.J.
Ward and T.J. Peters. Mössbauer spectroscopy, electron microscopy and
electron diffraction studies of the iron cores in various human and
animal haemosiderins, Biochim. Biophys. Acta. 957 (1988) 81-90.
- S. Mann, V.J. Wade, D.P.E. Dickson, N.M.K. Reid, R.J.
Ward, M. O'Connell and T.J. Peters. Structural specificity of
haemosiderin iron cores in iron-overload diseases, FEBS Lett. 234
(1988) 69-72.
- T.G. St. Pierre, W. Chua-anusorn, J. Webb, D. Macey, P.
Pootrakul, The form of iron oxide deposits in thalassemic tissues
varies between different groups of patients: a comparison between Thai
b-thalassemia/hemoglobin E patients and Australian b-thalassemia
patients, Biochim. Biophys. Acta 1407 (1998) 51-60.
- P. Gillis and S.H. Koenig. Transverse relaxation of solvent
protons induced by magnetized spheres: application to ferritin,
erythrocytes, and magnetite, Magn. Reson. Med. 5 (1987) 323-345.
- S.H. Koenig (1990) in Iron biominerals (R.B. Frankel, R.P. Blakemore, eds.), pp. 359-372, Plenum Press, New York.
- Allen, P.D., St Pierre, T.G., Chua-anusorn, W., Ström, V.,
and Rao, K.V. (2000) Low-frequency low-field magnetic susceptibility of
ferritin and hemosiderin. Biochim. Biophys. Acta, 1500, 186-196
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