(4,4′-diaponeurosporene), 2 lipids, and 1 [4Fe-4S]
cluster; four of these cofactors had not previously been described in the Protein Data Bank
(see fig. S2). The PshA and PshX polypeptides
account for ~140 kDa of the homodimeric RC.
The total molecular mass is ~200 kDa, owing to
the high cofactor content.
As a consequence of the homodimeric core,
the ET cofactors are arranged into two identical
branches about the C2 symmetry axis (Fig. 3).
The first ET cofactor (P800) is a pair of BChl g′
molecules, the stereoisomer at the 132 position
of ring E of BChl g (20), and is located toward
the positive side (P-side, equivalent to the outside)
of the membrane. On the negative side (N-side,
equivalent to the cytoplasm), the segment between
TMH8 and 9 of each PshA has two conserved Cys
residues that ligate FX, the terminal [4Fe-4S]
cluster. Between P800 and FX on each symmetric
branch are a BChl g and an 81-OH–Chl aF.
The antenna BChls form two layers within the
RC (dotted gray boxes in fig. S3A). Most are
relatively close to one another (i.e., within 6 Å),
which would allow rapid excitation-energy transfer. The bulk of this population (blue in Fig. 2B)
is ≥14 Å away from the ET chain (red in Fig. 2B).
There are only three antenna pigments on each
side that are within 13 Å of any BChls and
Chl in the ET chain (teal in Fig. 2B). Energy
transferred to the ET chain from the antenna
pool probably arrives via one of these six BChl g
Most of the antenna BChls (50 of 54) are
coordinated by the two PshA polypeptides, primarily from the six TMHs of the antenna domain
(Figs. 1 and 2); the other four BChl g are coordinated by the PshX subunits. Of the 27 BChls and
1 Chl associated with each PshA polypeptide, 19
are coordinated by His side chains, 1 by Asn, 1 by
Glu, 1 by Gln, 5 by water molecules, and 1 by an
unidentified atom that has higher electron density than a water molecule, hypothetically a
chloride ion (purple sphere in Figs. 3 and 4B).
Both BChls associated with PshX are His coordinated. Surprisingly, there is a BChl g′ associated
with each PshA within the center of an antenna
pigment cluster (green in fig. S3B).
A notable feature of the ET chain is that it con-
tains three chemically distinct BChls. It begins
on the P-side of the HbRC with P800, a BChl g′
dimer that serves as the primary electron donor,
and it ends on the N-side with FX, a [4Fe-4S]
cluster that serves as the terminal acceptor
(Fig. 3). Each BChl g′ in P800 is coordinated by
His537 from TMH10, whereas FX is ligated by
Cys432 and Cys441 from the loop between TMH8
and 9 of each PshA. The primary donor (P800)
and terminal acceptor (FX) are located on the C2
symmetry axis. There are two pairs of BChls on
each side of the symmetry axis that link P800 to
FX. The one closer to P800 is a BChl g, whereas the
one closer to FX is an 81-OH–Chl aF, in agreement
with modeling of the electrochromic bandshift of
the 81-OH–Chl aF spectrum in the presence of
oxidized P800+ (21).
On the basis of biochemical and biophysical
measurements (21–23), it was expected that the
primary acceptor in the HbRC, called A0 by analogy with the corresponding cofactor in PSI, was
an 81-OH–Chl aF molecule. The cofactor between
the special pair and primary acceptor is often
referred to as the “accessory” BChl and Chl (Acc).
We will use that term here, but it should not be
taken as a functional designation, as the analogous cofactor has been suggested to serve as the
primary electron donor in PSI (24, 25), PSII
(26), and the PbRC (27–29). Acc is coordinated by
a small molecule approximately the size of water,
and A0 is coordinated by a water molecule. These
are H bonded by the side chains of Gln458 and
Ser545, respectively (see discussion later in text).
As in the other RCs, within a branch of the ET
chain, the special pair (P800) and primary acceptor (A0) are coordinated by one polypeptide,
whereas the accessory BChl is coordinated by
the other polypeptide of the dimer.
The distances between cofactors (table S2) can
be used to explain the differences observed in ET
The role of the menaquinone (MQ) in the
HbRC has been controversial. The HbRC that
was crystallized in this study lacked MQ (fig.
S4), as it is not bound tightly by the protein and
thus is not present in this structure. However,
the edge-to-edge distance between A0 and FX is
Fig. 1. Overall structure of the HbRC. Structure as viewed from (A) the N-side or (B) within the
membrane. The two PshA polypeptides are colored in red and pink. PshX subunits are colored in orange.
Cofactor molecules are shown as stick models and colored teal (ET), blue (antenna), and lime
(carotenoids). The [4Fe-4S] cluster is shown as red (Fe) and yellow (S) spheres. BChl and Chl tails have
been truncated for clarity.
Fig. 2. Arrangement of the TMHs and pigments in the HbRC compared with PSI. (A) TMH
arrangement of the HbRC and its superposition with the TMHs of PSI (N-side view). TMHs of
the two PshA subunits are colored in red and pink and labeled 1 to 11, from N to C terminus. PshX helices
are colored in orange and are labeled “X.” Transparent gray helices are from the heterodimeric core
of PSI [Protein Data Bank (PDB) ID 1JB0]. (B) N-side view of the cofactor organization in the HbRC
(colored) superimposed with the cofactors associated with the PsaA-PsaB heterodimeric core of PSI
(transparent gray, PDB ID 1JB0). ET BChls and Chls are colored red, bulk antenna pigments are colored
blue, and the three antenna BChl g that flank the ETchain are colored teal. BChl and Chl tails have
been truncated for clarity.