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A study of the ultrastructure of fragile-X-related proteins.

Sjekloća L, Konarev PV, Eccleston J, Taylor IA, Svergun DI, Pastore A - Biochem. J. (2009)

Bottom Line: In the present study, we describe how we have produced overlapping recombinant fragments of human FMRP and its paralogues which encompass the evolutionary conserved region.We have studied their behaviour in solution by complementary biochemical and biophysical techniques, identified the regions which promote self-association and determined their overall three-dimensional shape.The present study paves the way to further studies and rationalizes the existing knowledge on the self-association properties of these proteins.

View Article: PubMed Central - PubMed

Affiliation: National Institute for Medical Research, The Ridgeway, London NW7 1AA, UK.

ABSTRACT
Fragile-X-related proteins form a family implicated in RNA metabolism. Their sequence is composed of conserved N-terminal and central regions which contain Tudor and KH domains and of a divergent C-terminus with motifs rich in arginine and glycine residues. The most widely studied member of the family is probably FMRP (fragile X mental retardation protein), since absence or mutation of this protein in humans causes fragile X syndrome, the most common cause of inherited mental retardation. Understanding the structural properties of FMRP is essential for correlating it with its functions. The structures of isolated domains of FMRP have been reported, but nothing is yet known with regard to the spatial arrangement of the different modules, partly because of difficulties in producing both the full-length protein and its multidomain fragments in quantities, purities and monodispersity amenable for structural studies. In the present study, we describe how we have produced overlapping recombinant fragments of human FMRP and its paralogues which encompass the evolutionary conserved region. We have studied their behaviour in solution by complementary biochemical and biophysical techniques, identified the regions which promote self-association and determined their overall three-dimensional shape. The present study paves the way to further studies and rationalizes the existing knowledge on the self-association properties of these proteins.

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Choice and production of the multidomain constructs used in the present study(A) Schematic representation of the architecture of FXR proteins. The different sequence motifs are indicated with different geometric shapes. (B) Summary of the constructs produced and their properties. N.P., not produced.
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Figure 1: Choice and production of the multidomain constructs used in the present study(A) Schematic representation of the architecture of FXR proteins. The different sequence motifs are indicated with different geometric shapes. (B) Summary of the constructs produced and their properties. N.P., not produced.

Mentions: With the ultimate goal to fully characterize the FXR proteins, we systematically produced overlapping constructs for all three human homologues covering two or more tandem domains (Figure 1). The construct boundaries were chosen according to the specific motif consensus sequences and to the multiple alignment of the FXR family (Supplementary Figure S1). All expressed constructs resulted in the soluble fraction, but had very different behaviours towards degradation and solubility.


A study of the ultrastructure of fragile-X-related proteins.

Sjekloća L, Konarev PV, Eccleston J, Taylor IA, Svergun DI, Pastore A - Biochem. J. (2009)

Choice and production of the multidomain constructs used in the present study(A) Schematic representation of the architecture of FXR proteins. The different sequence motifs are indicated with different geometric shapes. (B) Summary of the constructs produced and their properties. N.P., not produced.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2662490&req=5

Figure 1: Choice and production of the multidomain constructs used in the present study(A) Schematic representation of the architecture of FXR proteins. The different sequence motifs are indicated with different geometric shapes. (B) Summary of the constructs produced and their properties. N.P., not produced.
Mentions: With the ultimate goal to fully characterize the FXR proteins, we systematically produced overlapping constructs for all three human homologues covering two or more tandem domains (Figure 1). The construct boundaries were chosen according to the specific motif consensus sequences and to the multiple alignment of the FXR family (Supplementary Figure S1). All expressed constructs resulted in the soluble fraction, but had very different behaviours towards degradation and solubility.

Bottom Line: In the present study, we describe how we have produced overlapping recombinant fragments of human FMRP and its paralogues which encompass the evolutionary conserved region.We have studied their behaviour in solution by complementary biochemical and biophysical techniques, identified the regions which promote self-association and determined their overall three-dimensional shape.The present study paves the way to further studies and rationalizes the existing knowledge on the self-association properties of these proteins.

View Article: PubMed Central - PubMed

Affiliation: National Institute for Medical Research, The Ridgeway, London NW7 1AA, UK.

ABSTRACT
Fragile-X-related proteins form a family implicated in RNA metabolism. Their sequence is composed of conserved N-terminal and central regions which contain Tudor and KH domains and of a divergent C-terminus with motifs rich in arginine and glycine residues. The most widely studied member of the family is probably FMRP (fragile X mental retardation protein), since absence or mutation of this protein in humans causes fragile X syndrome, the most common cause of inherited mental retardation. Understanding the structural properties of FMRP is essential for correlating it with its functions. The structures of isolated domains of FMRP have been reported, but nothing is yet known with regard to the spatial arrangement of the different modules, partly because of difficulties in producing both the full-length protein and its multidomain fragments in quantities, purities and monodispersity amenable for structural studies. In the present study, we describe how we have produced overlapping recombinant fragments of human FMRP and its paralogues which encompass the evolutionary conserved region. We have studied their behaviour in solution by complementary biochemical and biophysical techniques, identified the regions which promote self-association and determined their overall three-dimensional shape. The present study paves the way to further studies and rationalizes the existing knowledge on the self-association properties of these proteins.

Show MeSH
Related in: MedlinePlus