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PDBsum entry 2ezh
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DNA binding protein
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PDB id
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2ezh
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References listed in PDB file
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Key reference
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Title
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Solution structure of the i gamma subdomain of the mu end DNA-Binding domain of phage mu transposase.
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Authors
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R.T.Clubb,
S.Schumacher,
K.Mizuuchi,
A.M.Gronenborn,
G.M.Clore.
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Ref.
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J Mol Biol, 1997,
273,
19-25.
[DOI no: ]
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PubMed id
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Abstract
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The MuA transposase of phase Mu is a large modular protein that plays a central
role in transposition. We show that the Mu end DNA-binding domain, I beta gamma,
which is responsible for binding the DNA attachment sites at each end of the Mu
genome, comprises two subdomains, I beta and I gamma, that are structurally
autonomous and do not interact with each other in the absence of DNA. The
solution structure of the I gamma subdomain has been determined by
multidimensional NMR spectroscopy. The structure of I gamma comprises a four
helix bundle and, despite the absence of any significant sequence identity, the
topology of the first three helices is very similar to that of the homeodomain
family of helix-turn-helix DNA-binding proteins. The helix-turn-helix motif of I
gamma, however, differs from that of the homeodomains in so far as the loop is
longer and the second helix is shorter, reminiscent of that in the POU-specific
domain.
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Figure 1.
Figure 1. a, A drawing of the domain organization of
MuA transposase, and of the various fragments studied
by NMR. b, Sequence of Ig (residues 174 to 247) with
the location of the four helices determined by NMR
indicated. MuA deletion variants were constructed
using PCR methodology. Plasmid pMK609 (Mizuuchi &
Mizuuchi, 1989), which contains DNA encoding the
MuA transposase, was targeted for deletion. Appropri-
ate restriction sites were engineered into the PCR pri-
mers allowing selectively amplified DNAs to be ligated
into the phage T7 RNA polymerase expression plasmid
pET3c (Novagen). The expression plasmids were then
transferred to E. coli strain BL21(DE3) for protein
expression. Three deletion constructs of the MuA trans-
posase were constructed and comprise residues 77 to
247 (Ibg), residues 77 to 174 (Ib), and residues 174 to
247 (Ig) of MuA transposase.
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Figure 4.
Figure 4. Comparison of the Ig domain (a) with the Oct-1 POU homeodomain (b). The two structures can be super-
imposed with a C
a
atomic rms difference of 2 Å for 41 residues (residues 179 to 194, 199 to 212 and 223 to 233 of Ig and
residues 107 to 122, 126 to 139 and 147 to 157 of the Oct-1 POU homeodomain) comprising principally helices 1, 2 and 3.
Helices 1, 2 and 3 of Ig comprise residues 182 to 192, 200 to 214 and 221 to 231, respectively, while in the recognition
helix of the POU homedomain these three helices extend from residues 110 to 122, 128 to 137 and 142 to 160, respect-
ively. The HTH motif is shown in yellow, and the other helices in red. The coordinates of the Oct-1 POU homeodomain
are taken from Klemm et al. (1994). The Figure was generated with the program MOLMOL (Konradi et al., 1996).
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The above figures are
reprinted
by permission from Elsevier:
J Mol Biol
(1997,
273,
19-25)
copyright 1997.
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