D-family DNA polymerase - DP2
subunit
Julia Borys '21 and Sam Schaffner '21
Contents:
I. Introduction
DNA polymerases (DNAPs) have evolved across all life forms in
order to perform highly accurate and processive replication of the
genome, repair of DNA, and resolution of Okazaki fragments. DNA
polymerase D (PolD) is an atypical class of DNA polymerases found in
archaea. PolD initiates DNA synthesis for both the leading and lagging
strands in all archaea with the exception of
Crenarchaea. PolD contains
two subunits: a proofreading exonuclease subunit (DP1) and a polymerase
catalytic subunit (DP2), shown here from hyperthermophilic archaea
species
Pyrococcus abyssi. The active site of DP2 shows
surprising structural similarity to the “two-double-psi ß-barrel” (DPBB)
family of RNAPs, suggesting a complex evolutionary history to the
archaeal DNA polymerase.
II. Structure of DP2
The polymerase catalytic DP2 subunit of PolD is made of
the N-terminal domain, the central domain, the catalytic
domain, and part of the C-terminal domain. There is substantial
interaction between the N-terminal domain and the catalytic domain
that forms the DNAP active site surrounded by two double-psi
ß-barrels (DPBBs). The N-terminal domain has a 45-residue long
N-terminal extension named the
, which interacts with the catalytic domain (ß33). The
N-terminal self-assembly region has a long
that makes the most connections with the catalytic domain
and a short
. The interactions between the N-terminal domain and catalytic domain include
,
contacts between
hydrophobic residues, and secondary structure interactions between ß1
and ß33.
The DP2 subunit contains two
domains that sit on either side of
the protein’s
. This loop is solvent-exposed and located
within a crevice that is 55 angstroms long and 25 angstroms wide,
which is suited for a 1.5 helical turn of B-DNA.
The putative DNA binding region is located in the catalytic domain
between two zinc-modules,
Other than the tetrahedral coordination of the Zn2+
ions by two pairs of cysteines, the modules have separate structures.
III. DPBB Architecture
is located in the central domain. The crystallized
structure for DPBB only shows 5 of the 6 expected beta-strands, but the
missing strand is expected to be located in the stretch of 17 missing
residues. The central domain also contains a seven-stranded antiparallel
ß-sheet (ß10-12 and ß14-17) that is wrapped by six alpha-helices
(alpha13-14 and alpha19-22).
The catalytic domain has a central six-stranded
. The most conserved sequence motif of the DP2 is the
right-handed alpha-helix and a 10-residue long loop that connects the
fifth (ß31) and sixth (ß32) ß-strands. This motif contains the
catalytically critical aspartic residues
(also named D961 and D963 in reference to the
literature).
IV. Structural Relationship to Other RNAPs
The unique two DPBB motif structure of DP2 is not structurally
similar to any other known DNAPs. However, the two DPBB structure
shares unexpected homology to the ‘two-barrel’ family of RNAP. DBPP-A
of multi-subunit RNAP chelate catalytic Mg2+ with a DFDGDE signature.
The two invariant aspartic residues on DBPP-2 (D956 and D958) are
aligned with three catalytic aspartic residues in RNAP (see figure
below). DBPP-B of RNAP contains two lysine residues for DNA binding.
Unfortunately, the corresponding residues for PolD are found in a
stretch of missing residues.
VI. References
Raia P, Carroni M, Henry E, Pehau-Arnaudet G, Brûlé S,
Béguin P, et al. (2019). Structure of the DP1–DP2 PolD complex bound
with DNA and its implications for the evolutionary history of DNA and
RNA polymerases. PLoS Biol 17 (1), 3000122.
Sauguet L, Raia P, Henneke G et al. (2016). Shared
active site architecture between archaeal PolD and multi-subunit RNA
polymerases revealed by X-ray crystallography. Nat Commun 7,
12227.
Shen Y et al. (2001). Invariant Asp-1122 and Asp-1124
are essential residues for polymerization catalysis of family D DNA
polymerase from Pyrococcus horikoshii. J. Biol. Chem. 276,
27376–27383.
Werner F, Grohmann D. (2011). Evolution of
multisubunit RNA polymerases in the three domains of life. Nat.
Rev. Microbiol. 9, 85–98.
Amino
Acid Color Key
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