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Conformational rearrangement of gastric H(+),K(+)-ATPase induced by an acid suppressant.

Abe K, Tani K, Fujiyoshi Y - Nat Commun (2011)

Bottom Line: The density of the bound SCH28080 is found near transmembrane (TM) helices 4, 5 and 6, in the luminal cavity.The SCH28080-binding site is formed by the rearrangement of TM helices, which is in turn transmitted to the cytoplasmic domains, resulting in a luminal-open conformation.These results represent the first structural evidence for a binding site of an acid suppressant on H(+),K(+)-ATPase, and the conformational change induced by this class of drugs.

View Article: PubMed Central - PubMed

Affiliation: Department of Biophysics, Faculty of Science, Kyoto University, Oiwake, Kitashirakawa, Sakyo-ku, Kyoto 606-0852, Japan.

ABSTRACT
Acid-related gastric diseases are associated with disorder of digestive tract acidification. The gastric proton pump, H(+),K(+)-ATPase, exports H(+) in exchange for luminal K(+) to generate a highly acidic environment in the stomach, and is a main target for acid suppressants. Here, we report the three-dimensional structure of gastric H(+),K(+)-ATPase with bound SCH28080, a representative K(+)-competitive acid blocker, at 7 Å resolution based on electron crystallography of two-dimensional crystals. The density of the bound SCH28080 is found near transmembrane (TM) helices 4, 5 and 6, in the luminal cavity. The SCH28080-binding site is formed by the rearrangement of TM helices, which is in turn transmitted to the cytoplasmic domains, resulting in a luminal-open conformation. These results represent the first structural evidence for a binding site of an acid suppressant on H(+),K(+)-ATPase, and the conformational change induced by this class of drugs.

No MeSH data available.


Related in: MedlinePlus

Rearrangement of TM helices induced by SCH28080 binding.(a) Comparison of the TM helices between E2AlF and (SCH)E2BeF structures. A cross-section of the luminal TM region parallel to the membrane plane is viewed from the luminal side of the membrane. Mesh represents the EM density map (1σ) of (SCH)E2BeF, with homology models of (SCH)E2BeF (colour ribbons as in Fig. 1a) and E2AlF (grey ribbon)14 superimposed. A cross-section parallel to the membrane plane displays the contour level at the indicated plane (colour gradually changes from blue (1σ) to red (5σ) as indicated in the figure). White arrows indicate the conformational change of M1–M4 induced by the SCH28080 binding (red stick as in Fig. 1b). The position of the slice is indicated as a red box in Supplementary Fig. S4. (b) A view of the M1–M4 region perpendicular to the membrane plane showing inwardly swinging movement of the M1–M2 helix bundle (blue ribbon) along with an outwards movement of M3–M4 (cyan ribbon) compared with the E2AlF homology model (grey ribbon). Dotted lines indicate the approximate position of the lipid bilayer.
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f3: Rearrangement of TM helices induced by SCH28080 binding.(a) Comparison of the TM helices between E2AlF and (SCH)E2BeF structures. A cross-section of the luminal TM region parallel to the membrane plane is viewed from the luminal side of the membrane. Mesh represents the EM density map (1σ) of (SCH)E2BeF, with homology models of (SCH)E2BeF (colour ribbons as in Fig. 1a) and E2AlF (grey ribbon)14 superimposed. A cross-section parallel to the membrane plane displays the contour level at the indicated plane (colour gradually changes from blue (1σ) to red (5σ) as indicated in the figure). White arrows indicate the conformational change of M1–M4 induced by the SCH28080 binding (red stick as in Fig. 1b). The position of the slice is indicated as a red box in Supplementary Fig. S4. (b) A view of the M1–M4 region perpendicular to the membrane plane showing inwardly swinging movement of the M1–M2 helix bundle (blue ribbon) along with an outwards movement of M3–M4 (cyan ribbon) compared with the E2AlF homology model (grey ribbon). Dotted lines indicate the approximate position of the lipid bilayer.

Mentions: Comparison between the E2P* and (SCH)E2BeF structures of H+,K+-ATPase reveals that binding of SCH28080 induces the rearrangement of, mostly, the M1–M4 helices at the luminal side of the enzyme (Fig. 3). Although all TM helices are located in similar positions on the cytoplasmic side of the membrane (Supplementary Fig. S4a,b), M3 and M4 bend outwardly at the luminal membrane interface (Fig. 3a, Supplementary Fig. S4c). This difference, which is also observed in the ouabain-bound Na+,K+-ATPase structure20, clears space at the luminal cavity to create the binding site for SCH28080, where M4 locates in the H+,K+-ATPase E2P* structure (Fig. 3a). At the same time, the helix bundle of M1 and M2 moves towards the cytoplasmic side along with an outwards movement of M4 (Fig. 3). As a consequence, M1 and M2 come closer and apparently restrict the SCH28080-binding site (Fig. 3a), which is consistent with the slow dissociation of SCH28080 (ref. 16), the fourfold lower affinity of L141C mutant located in M2 (ref. 28) (Fig. 1b,c), and the photoaffinity labelling between the M1–M2 segment and a SCH28080 derivative22. Such an inwards swinging motion of the M1–M2 bundle (Fig. 3a,b) has not been observed in the Na+,K+-ATPase structure with bound ouabain in the low-affinity mode (Supplementary Fig. S5), but has been predicted in the high-affinity ouabain-binding model based on the SERCA E2BeF structure2021, which is consistent with the previously reported important contribution of the M1–M2 segment to the high-affinity ouabain binding33. Therefore, the M1–M2 movement observed in the H+,K+-ATPase structure (Supplementary Fig. S5) seems to be commonly involved in the formation of the high-affinity inhibitor-binding pocket on both closely related ATPases. Overall, this rearrangement of the TM helices of the H+,K+-ATPase results in the luminal-open conformation, similar to that observed in the SERCA E2BeF structure2130, which presumably allows for the release of bound Ca2+.


Conformational rearrangement of gastric H(+),K(+)-ATPase induced by an acid suppressant.

Abe K, Tani K, Fujiyoshi Y - Nat Commun (2011)

Rearrangement of TM helices induced by SCH28080 binding.(a) Comparison of the TM helices between E2AlF and (SCH)E2BeF structures. A cross-section of the luminal TM region parallel to the membrane plane is viewed from the luminal side of the membrane. Mesh represents the EM density map (1σ) of (SCH)E2BeF, with homology models of (SCH)E2BeF (colour ribbons as in Fig. 1a) and E2AlF (grey ribbon)14 superimposed. A cross-section parallel to the membrane plane displays the contour level at the indicated plane (colour gradually changes from blue (1σ) to red (5σ) as indicated in the figure). White arrows indicate the conformational change of M1–M4 induced by the SCH28080 binding (red stick as in Fig. 1b). The position of the slice is indicated as a red box in Supplementary Fig. S4. (b) A view of the M1–M4 region perpendicular to the membrane plane showing inwardly swinging movement of the M1–M2 helix bundle (blue ribbon) along with an outwards movement of M3–M4 (cyan ribbon) compared with the E2AlF homology model (grey ribbon). Dotted lines indicate the approximate position of the lipid bilayer.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
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f3: Rearrangement of TM helices induced by SCH28080 binding.(a) Comparison of the TM helices between E2AlF and (SCH)E2BeF structures. A cross-section of the luminal TM region parallel to the membrane plane is viewed from the luminal side of the membrane. Mesh represents the EM density map (1σ) of (SCH)E2BeF, with homology models of (SCH)E2BeF (colour ribbons as in Fig. 1a) and E2AlF (grey ribbon)14 superimposed. A cross-section parallel to the membrane plane displays the contour level at the indicated plane (colour gradually changes from blue (1σ) to red (5σ) as indicated in the figure). White arrows indicate the conformational change of M1–M4 induced by the SCH28080 binding (red stick as in Fig. 1b). The position of the slice is indicated as a red box in Supplementary Fig. S4. (b) A view of the M1–M4 region perpendicular to the membrane plane showing inwardly swinging movement of the M1–M2 helix bundle (blue ribbon) along with an outwards movement of M3–M4 (cyan ribbon) compared with the E2AlF homology model (grey ribbon). Dotted lines indicate the approximate position of the lipid bilayer.
Mentions: Comparison between the E2P* and (SCH)E2BeF structures of H+,K+-ATPase reveals that binding of SCH28080 induces the rearrangement of, mostly, the M1–M4 helices at the luminal side of the enzyme (Fig. 3). Although all TM helices are located in similar positions on the cytoplasmic side of the membrane (Supplementary Fig. S4a,b), M3 and M4 bend outwardly at the luminal membrane interface (Fig. 3a, Supplementary Fig. S4c). This difference, which is also observed in the ouabain-bound Na+,K+-ATPase structure20, clears space at the luminal cavity to create the binding site for SCH28080, where M4 locates in the H+,K+-ATPase E2P* structure (Fig. 3a). At the same time, the helix bundle of M1 and M2 moves towards the cytoplasmic side along with an outwards movement of M4 (Fig. 3). As a consequence, M1 and M2 come closer and apparently restrict the SCH28080-binding site (Fig. 3a), which is consistent with the slow dissociation of SCH28080 (ref. 16), the fourfold lower affinity of L141C mutant located in M2 (ref. 28) (Fig. 1b,c), and the photoaffinity labelling between the M1–M2 segment and a SCH28080 derivative22. Such an inwards swinging motion of the M1–M2 bundle (Fig. 3a,b) has not been observed in the Na+,K+-ATPase structure with bound ouabain in the low-affinity mode (Supplementary Fig. S5), but has been predicted in the high-affinity ouabain-binding model based on the SERCA E2BeF structure2021, which is consistent with the previously reported important contribution of the M1–M2 segment to the high-affinity ouabain binding33. Therefore, the M1–M2 movement observed in the H+,K+-ATPase structure (Supplementary Fig. S5) seems to be commonly involved in the formation of the high-affinity inhibitor-binding pocket on both closely related ATPases. Overall, this rearrangement of the TM helices of the H+,K+-ATPase results in the luminal-open conformation, similar to that observed in the SERCA E2BeF structure2130, which presumably allows for the release of bound Ca2+.

Bottom Line: The density of the bound SCH28080 is found near transmembrane (TM) helices 4, 5 and 6, in the luminal cavity.The SCH28080-binding site is formed by the rearrangement of TM helices, which is in turn transmitted to the cytoplasmic domains, resulting in a luminal-open conformation.These results represent the first structural evidence for a binding site of an acid suppressant on H(+),K(+)-ATPase, and the conformational change induced by this class of drugs.

View Article: PubMed Central - PubMed

Affiliation: Department of Biophysics, Faculty of Science, Kyoto University, Oiwake, Kitashirakawa, Sakyo-ku, Kyoto 606-0852, Japan.

ABSTRACT
Acid-related gastric diseases are associated with disorder of digestive tract acidification. The gastric proton pump, H(+),K(+)-ATPase, exports H(+) in exchange for luminal K(+) to generate a highly acidic environment in the stomach, and is a main target for acid suppressants. Here, we report the three-dimensional structure of gastric H(+),K(+)-ATPase with bound SCH28080, a representative K(+)-competitive acid blocker, at 7 Å resolution based on electron crystallography of two-dimensional crystals. The density of the bound SCH28080 is found near transmembrane (TM) helices 4, 5 and 6, in the luminal cavity. The SCH28080-binding site is formed by the rearrangement of TM helices, which is in turn transmitted to the cytoplasmic domains, resulting in a luminal-open conformation. These results represent the first structural evidence for a binding site of an acid suppressant on H(+),K(+)-ATPase, and the conformational change induced by this class of drugs.

No MeSH data available.


Related in: MedlinePlus