which partially neutralize acidic residues at the binding sites. In this process, the pump switches back
to the calcium-free E2 state ④. This is followed by closure of the luminal channel and occlusion of the
bound protons ⑤. In the last step ⑥, Asp351 is dephosphorylated, leading to the reformation of H n E2:
ATP.
Even though neither the structure nor the function of the nAChR (§ 1.6.2) and SERCA has
anything in common, it is worth pointing out some similarities and differences in the way each MP
couples the conformations of its TM and extramembrane regions. In the case of the nAChR, the
binding of acetylcholine (ACh) initiates a rotational rearrangement of the extracellular domains, which
affects the position of TM helix M2, leading to the opening of the TM ion channel, more than 60 Å
away. In the case of SERCA, the binding of cytosolic Ca
2+ ions to TM sites induces a rearrangement of
the TM helix bundle, as a result of which extramembrane domains are forced to move with respect to
each other, initiating the phosphorylation/dephosphorylation cycle that powers calcium pumping.
Movements of the cytosolic domains in the course of this cycle, in turn, entail a further rearrangement
of the TM helices, imposed by the links connecting cytosolic domain A to the M1–M3 helices, which
results in the Ca
2+ ions being released into the SR lumen. Whereas TM rearrangements are much larger
in SERCA than in pLGICs (compare Figs. 1.36 and 1.30), a key feature of the two protein families is
the coupling of TM to extramembrane conformational changes. Such is not the case with BR, in which
all key events occur within the TM region (§ 1.6.1). In visual rhodopsin, however, which shares the
same cofactor, the helix movements initiated by the light-driven isomerization of retinal result not in
proton pumping but in a rearrangement of the cytosolic region. This is detected by soluble proteins,
which relay and amplify the signal. The same mechanism is exploited by class A (rhodopsin-like)
GPCRs, whose cytosolic conformation changes as a result of the binding of extracellular ligands to a
cleft formed by the TM helices, thus achieving TM signaling (see Chap. 2, § 2.5.2).
We have noted above that the two TM calcium-binding sites of SERCA are formed by residues
carried by helices M4, M5, M6, and M8 (Toyoshima et al. 2000; Obara et al. 2005) (Fig. 1.35). As a
result, SERCA1a is strongly stabilized by Ca
2+ against thermal- and detergent-induced denaturation
(Møller et al. 1980; Merino et al. 1994), presumably because the bridging of TM helices by Ca
2+ ions
opposes opening of the TM domain. This observation is of great interest when trying to understand the
mechanism by which APols stabilize the calcium pump and, presumably, other MPs (Chap. 5, § 5.6)
A molecule of PE is bound in a cavity between TM helices M2 and M4 in the calcium-free
conformation of the protein, acting as a wedge keeping these helices apart (Obara et al. 2005)
(Fig. 1.37). In the calcium-bound form, the cavity is closed, so that the PE molecule must be displaced
for Ca
2+ to bind (Obara et al. 2005). As noted by A.G. Lee (2011a, b), this could possibly provide an
explanation for some of the effects of PE on Ca
2+ -ATPase function (Starling et al. 1996). It may also
perhaps be related to the high sensitivity to detergents exhibited by SERCA1a in its calcium-free form
(see Chap. 5, § 5.6.1). A detailed analysis of this and other lipid-binding sites on SERCA1a is given by
Drachmann et al. (2014).
In a recent development, contrast variation has permitted Chikashi Toyoshima and his
colleagues to visualize the important rocking movements of SERCA1a with respect to the surrounding
lipids and the rearrangement of molecular interactions, involving particularly tryptophan and basic
residues that accompany the enzymatic cycle. These movements permit the hydrophobic surface of the
TM domain to remain lipid-buried throughout (see Norimatsu et al. 2017; the online version of the
article includes impressive videos of the rearrangements the protein and lipids undergo in the course of
the cycle; see also comments in Sweadner 2017).
1.6 Dynamics of Transmembrane Regions and the Function of Membrane Proteins
47
to the calcium-free E2 state ④. This is followed by closure of the luminal channel and occlusion of the
bound protons ⑤. In the last step ⑥, Asp351 is dephosphorylated, leading to the reformation of H n E2:
ATP.
Even though neither the structure nor the function of the nAChR (§ 1.6.2) and SERCA has
anything in common, it is worth pointing out some similarities and differences in the way each MP
couples the conformations of its TM and extramembrane regions. In the case of the nAChR, the
binding of acetylcholine (ACh) initiates a rotational rearrangement of the extracellular domains, which
affects the position of TM helix M2, leading to the opening of the TM ion channel, more than 60 Å
away. In the case of SERCA, the binding of cytosolic Ca
2+ ions to TM sites induces a rearrangement of
the TM helix bundle, as a result of which extramembrane domains are forced to move with respect to
each other, initiating the phosphorylation/dephosphorylation cycle that powers calcium pumping.
Movements of the cytosolic domains in the course of this cycle, in turn, entail a further rearrangement
of the TM helices, imposed by the links connecting cytosolic domain A to the M1–M3 helices, which
results in the Ca
2+ ions being released into the SR lumen. Whereas TM rearrangements are much larger
in SERCA than in pLGICs (compare Figs. 1.36 and 1.30), a key feature of the two protein families is
the coupling of TM to extramembrane conformational changes. Such is not the case with BR, in which
all key events occur within the TM region (§ 1.6.1). In visual rhodopsin, however, which shares the
same cofactor, the helix movements initiated by the light-driven isomerization of retinal result not in
proton pumping but in a rearrangement of the cytosolic region. This is detected by soluble proteins,
which relay and amplify the signal. The same mechanism is exploited by class A (rhodopsin-like)
GPCRs, whose cytosolic conformation changes as a result of the binding of extracellular ligands to a
cleft formed by the TM helices, thus achieving TM signaling (see Chap. 2, § 2.5.2).
We have noted above that the two TM calcium-binding sites of SERCA are formed by residues
carried by helices M4, M5, M6, and M8 (Toyoshima et al. 2000; Obara et al. 2005) (Fig. 1.35). As a
result, SERCA1a is strongly stabilized by Ca
2+ against thermal- and detergent-induced denaturation
(Møller et al. 1980; Merino et al. 1994), presumably because the bridging of TM helices by Ca
2+ ions
opposes opening of the TM domain. This observation is of great interest when trying to understand the
mechanism by which APols stabilize the calcium pump and, presumably, other MPs (Chap. 5, § 5.6)
A molecule of PE is bound in a cavity between TM helices M2 and M4 in the calcium-free
conformation of the protein, acting as a wedge keeping these helices apart (Obara et al. 2005)
(Fig. 1.37). In the calcium-bound form, the cavity is closed, so that the PE molecule must be displaced
for Ca
2+ to bind (Obara et al. 2005). As noted by A.G. Lee (2011a, b), this could possibly provide an
explanation for some of the effects of PE on Ca
2+ -ATPase function (Starling et al. 1996). It may also
perhaps be related to the high sensitivity to detergents exhibited by SERCA1a in its calcium-free form
(see Chap. 5, § 5.6.1). A detailed analysis of this and other lipid-binding sites on SERCA1a is given by
Drachmann et al. (2014).
In a recent development, contrast variation has permitted Chikashi Toyoshima and his
colleagues to visualize the important rocking movements of SERCA1a with respect to the surrounding
lipids and the rearrangement of molecular interactions, involving particularly tryptophan and basic
residues that accompany the enzymatic cycle. These movements permit the hydrophobic surface of the
TM domain to remain lipid-buried throughout (see Norimatsu et al. 2017; the online version of the
article includes impressive videos of the rearrangements the protein and lipids undergo in the course of
the cycle; see also comments in Sweadner 2017).
1.6 Dynamics of Transmembrane Regions and the Function of Membrane Proteins
47