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PDBsum entry 2a8t
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Translation,hydrolase
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PDB id
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2a8t
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References listed in PDB file
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Key reference
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Title
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Crystal structures of u8 snorna decapping nudix hydrolase, X29, And its metal and cap complexes.
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Authors
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J.N.Scarsdale,
B.A.Peculis,
H.T.Wright.
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Ref.
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Structure, 2006,
14,
331-343.
[DOI no: ]
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PubMed id
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Abstract
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X29, a 25 kDa Nudix hydrolase from Xenopus laevis that cleaves 5' caps from U8
snoRNA, crystallizes as a homodimeric apoenzyme. Manganese binds crystals of
apo-X29 to form holo-X29 only in the presence of nucleot(s)ide. Structural
changes in X29 on nucleo-t(s)ide-assisted Mn(+2) uptake account for the observed
cooperativity of metal binding. Structures of X29 with GTP or m7GpppA show a
different mode of ligand binding from that of other cap binding proteins and
suggest a possible three- or four-metal Nudix reaction mechanism. The X29 dimer
has no known RNA binding motif, but its striking surface dipolarity and unique
structural features create a plausible RNA binding channel on the positive face
of the protein. Because U8 snoRNP is essential for accumulation of mature 5.8S
and 28S rRNA in vertebrate ribosome biogenesis, and cap structures are required
for U8 stability in vivo, X29 could profoundly influence this fundamental
cellular pathway.
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Figure 3.
Figure 3. Surface Charge Distribution of Opposite Faces of
X29
Blue indicates regions of positive surface charge and
red indicates negative, contoured at ± 5kt cutoff, respectively.
Overlaid ribbon in (C) shows extensions (black arrow pointers)
that define the two sides of the inferred RNA binding channel,
which is indicated by yellow arrows. Active sites of each
monomer are occupied by m7GpppA (gold and green ball-and-stick).
Views are down the dyad axis.
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The above figure is
reprinted
by permission from Cell Press:
Structure
(2006,
14,
331-343)
copyright 2006.
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Secondary reference #1
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Title
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Crystals of X29, A xenopus laevis u8 snorna-Binding protein with nuclear decapping activity.
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Authors
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B.A.Peculis,
J.N.Scarsdale,
H.T.Wright.
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Ref.
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Acta Crystallogr D Biol Crystallogr, 2004,
60,
1668-1669.
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PubMed id
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Secondary reference #2
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Title
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Xenopus u8 snorna binding protein is a conserved nuclear decapping enzyme.
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Authors
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T.Ghosh,
B.Peterson,
N.Tomasevic,
B.A.Peculis.
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Ref.
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Mol Cell, 2004,
13,
817-828.
[DOI no: ]
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PubMed id
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Figure 1.
Figure 1. cDNA for X29 ProteinThe complete nucleotide
sequence for X29 is shown with a translation below. The
initiating methionine is indicated in bold. Underlined peptides
were obtained by Edman sequencing or MS/MS. The NUDIX consensus
sequence is boxed, with the adjacent glutamic acid residues
essential for catalysis indicated. The KK in the NUDIX domain
indicates the X29 EE→KK mutation (see text). Degenerate
oligonucleotides corresponding to peptides labeled #7 and #8
(dashed underline) yielded the 350 bp fragment by RT-PCR. X29
Genbank Accession Number, AY423379.
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Figure 6.
Figure 6. X29 Cleaves Capped RNAs and Leaves a 5′
Monophosphate(A) U8 or U3 RNAs were transcribed in the presence
of GpppG (G), m^7GpppG (m^7G), or m^227G pppG (TMG) cap analogs
as indicated. RNAs were incubated in a decapping reaction then
ligated to a radiolabeled tag RNA of 30 nt in length via a
bridge oligo (schematic at top, see text and Experimental
Procedures). Ligation products were resolved on a denaturing 8%
acrylamide gel. Autoradiography displayed only the ligated
products. Capped RNAs are not a substrate for ligation since a
5′ monophosphate is required.(B) Xenopus oocytes were
microinjected with 5 pMol-labeled U8 RNA synthesized with a
GpppG or no cap (pppG). At the time interval indicated (in
minutes), total RNA was isolated from the oocytes and resolved
on a denaturing acrylamide gel. Autoradiography demonstrates the
instability of uncapped U8 RNA in vivo.
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The above figures are
reproduced from the cited reference
with permission from Cell Press
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