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PDBsum entry 1qht
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* Residue conservation analysis
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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 structure of a pol alpha family DNA polymerase from the hyperthermophilic archaeon thermococcus sp. 9 degrees n-7.
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Authors
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A.C.Rodriguez,
H.W.Park,
C.Mao,
L.S.Beese.
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Ref.
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J Mol Biol, 2000,
299,
447-462.
[DOI no: ]
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PubMed id
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Abstract
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The 2.25 A resolution crystal structure of a pol alpha family (family B) DNA
polymerase from the hyperthermophilic marine archaeon Thermococcus sp. 9 degrees
N-7 (9 degrees N-7 pol) provides new insight into the mechanism of pol alpha
family polymerases that include essentially all of the eukaryotic replicative
and viral DNA polymerases. The structure is folded into NH(2)- terminal, editing
3'-5' exonuclease, and polymerase domains that are topologically similar to the
two other known pol alpha family structures (bacteriophage RB69 and the recently
determined Thermococcus gorgonarius), but differ in their relative orientation
and conformation.The 9 degrees N-7 polymerase domain structure is reminiscent of
the "closed" conformation characteristic of ternary complexes of the
pol I polymerase family obtained in the presence of their dNTP and DNA
substrates. In the apo-9 degrees N-7 structure, this conformation appears to be
stabilized by an ion pair. Thus far, the other apo-pol alpha structures that
have been determined adopt open conformations. These results therefore suggest
that the pol alpha polymerases undergo a series of conformational transitions
during the catalytic cycle similar to those proposed for the pol I family.
Furthermore, comparison of the orientations of the fingers and exonuclease
(sub)domains relative to the palm subdomain that contains the pol active site
suggests that the exonuclease domain and the fingers subdomain of the polymerase
can move as a unit and may do so as part of the catalytic cycle. This provides a
possible structural explanation for the interdependence of polymerization and
editing exonuclease activities unique to pol alpha family polymerases.We suggest
that the NH(2)-terminal domain of 9 degrees N-7 pol may be structurally related
to an RNA-binding motif, which appears to be conserved among archaeal
polymerases. The presence of such a putative RNA- binding domain suggests a
mechanism for the observed autoregulation of bacteriophage T4 DNA polymerase
synthesis by binding to its own mRNA. Furthermore, conservation of this domain
could indicate that such regulation of pol expression may be a characteristic of
archaea. Comparion of the 9 degrees N-7 pol structure to its mesostable homolog
from bacteriophage RB69 suggests that thermostability is achieved by shortening
loops, forming two disulfide bridges, and increasing electrostatic interactions
at subdomain interfaces.
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Figure 4.
Figure 4. Comparisons of 9°N-7 and RB69 pols in
different (sub)domains to indicate loop segments that are
shorter in 9°N-7 pol. Least-squares C^a superposition was
performed over the region in blue, and the domains were
separated for side-by-side comparison. Loop regions are shown in
magenta and their residue endpoints are labeled. (a) Comparison
of the exonuclease domains. Indicated with purple asterisks are
the active site carboxylates (mutated to Ala in the case of the
9°N-7exo - pol used in this study). (b) Comparison of the
palm domains. The three active-site carboxylate groups are
depicted with side-chains.
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Figure 5.
Figure 5. Least-squares C^a superpositions of 9°N-7 and
RB69 pols in the (a) palm subdomain or (b) exonuclease domain.
The 9°N-7 pol backbone is shown in yellow, and its
active-site carboxylate groups in gold. The RB69 pol backbone is
drawn in green, and its active-site residues in magenta. The
central b-sheet of the exonuclease domain is light blue
(9°N-7 pol) or dark blue (RB69 pol) to allow tracking of the
domain motion. The precise regions used in the palm and
exonuclease superpositions are shown in Figure 4. The
NH[2]-terminal domain has been omitted for clarity. Arrows in
(a) indicate the direction of fingers and exonuclease movement
when moving from (a) to (b).
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The above figures are
reprinted
by permission from Elsevier:
J Mol Biol
(2000,
299,
447-462)
copyright 2000.
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Secondary reference #1
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Title
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Crystallization and preliminary diffraction analysis of a hyperthermostable DNA polymerase from a thermococcus archaeon.
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Authors
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M.Zhou,
C.Mao,
A.C.Rodriguez,
J.R.Kiefer,
R.B.Kucera,
L.S.Beese.
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Ref.
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Acta Crystallogr D Biol Crystallogr, 1998,
54,
994-995.
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PubMed id
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