Crystal Structure of a Mammalian Voltage-Dependent
            <i>Shaker</i>
            Family K
            <sup>+</sup>
            Channel

Long, Stephen B.; Campbell, Ernest B.; MacKinnon, Roderick

doi:10.1126/science.1116269

Cited by 2,053 publications

(2,199 citation statements)

References 52 publications

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“…For the crystals of ICMT alone, two point mutations of surface residues were introduced to improve crystallizability (G151A and E154A). Transformation into P. pastoris , expression, and cryo-lysis were performed as previously described 27 .…”

Section: Methodsmentioning

confidence: 99%

Atomic structure of the eukaryotic intramembrane RAS methyltransferase ICMT

Diver

Pedi

Koide

et al. 2018

Nature

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The maturation of Ras GTPases, and ~200 other cellular CaaX proteins, involves three enzymatic steps: addition of a farnesyl or geranylgeranyl prenyl lipid to the cysteine (C) in the C-terminal CaaX motif, proteolytic cleavage of the aaX residues, and methylation of the exposed prenylcysteine residue at its terminal carboxylate1. This final step is catalyzed by isoprenylcysteine carboxyl methyltransferase (ICMT), a eukaryotic-specific integral membrane enzyme of the endoplasmic reticulum (ER)2. ICMT is the only cellular enzyme known to methylate prenylcysteine substrates; methylation is important for their biological functions, including the membrane localisations and subsequent activities of Ras1, prelamin A3, and Rab4. ICMT inhibition has potential for combating progeria3 and cancer5–8. Here we present an X-ray structure of ICMT, at 2.3 Å resolution, in complex with its cofactor, an ordered lipid molecule and a monobody inhibitor. The active site spans cytosolic and membrane-exposed regions, indicating distinct entry routes for its cytosolic methyl donor, S-adenosyl-L-methionine (AdoMet), and for prenylcysteine substrates, which are associated with the ER membrane. The structure suggests how ICMT overcomes the topographical challenge and unfavourable energetics of bringing two reactants that have different cellular localisations together in a membrane environment – a relatively uncharacterized, but defining feature of many integral membrane enzymes.

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Section: Methodsmentioning

confidence: 99%

Atomic structure of the eukaryotic intramembrane RAS methyltransferase ICMT

Diver

Pedi

Koide

et al. 2018

Nature

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“…In the case of the voltage-gated K + channel (Kv), electrophysiological studies with toxin blockers showed the functional unit to be a tetramer 11 , a finding that was confirmed by X-ray crystallography [12][13][14][15] . Assembly of the four subunits into the tetrameric channel forms the permeation pathway in the centre.…”

mentioning

confidence: 89%

The cytoplasmic coiled-coil mediates cooperative gating temperature sensitivity in the voltage-gated H+ channel Hv1

et al. 2012

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Hv1/VsoP is a dimeric voltage-gated H + channel in which the gating of one subunit is reportedly coupled to that of the other subunit within the dimer. The molecular basis for dimer formation and intersubunit coupling, however, remains unknown. Here we show that the carboxy terminus ends downstream of the s4 voltage-sensor helix twist in a dimer coiled-coil architecture, which mediates cooperative gating. We also show that the temperature-dependent activation of H + current through Hv1/VsoP is regulated by thermostability of the coiled-coil domain, and that this regulation is altered by mutation of the linker between s4 and the coiled-coil. Cooperative gating within the dimer is also dependent on the linker structure, which circular dichroism spectrum analysis suggests is α-helical. our results indicate that the cytoplasmic coiled-coil strands form continuous α-helices with s4 and mediate cooperative gating to adjust the range of temperatures over which Hv1/VsoP operates.

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“… a–i , Pairwise superposition of the pore domain in hTRPV6 with ( a ) rat TRPV1 18 (PDB ID: 5IRX; RMSD = 2.065 Å), ( b ) rabbit TRPV2 21 (PDB ID: 5AN8; RMSD = 3.757 Å), ( c ) rat TRPV2 24 (PDB ID: 5HI9; RMSD = 4.399 Å), ( d ) human TRPA1 23 (PDB ID: 3J9P; RMSD = 1.429 Å), ( e ) human PKD2 25 (PDB ID: 5T4D; RMSD = 2.676 Å), ( f ) KcsA from Streptomyces lividans 47 (PDB ID: 1BL8; RMSD = 2.708 Å), ( g ) MthK from M. thermautotrophicum 48 (PDB ID: 1LNQ; RMSD = 2.947 Å), ( h ) rat Shaker 49 (PDB ID: 2A79; RMSD = 2.487 Å) and ( i ) rat GluA2 AMPA-subtype iGluR 28 (PDB ID: 5WEO; RMSD = 2.044 Å). j , Sequence alignment for the pore region of human TRPV3–6, TRPA1 and PKD2, rat TRPV1, 2, 6, Shaker and GluA2, rabbit TRPV2 and bacterial K + channels KcsA and MthK.…”

Section: Extended Datamentioning

confidence: 99%

Opening of the human epithelial calcium channel TRPV6

McGoldrick

Singh

Saotome

et al. 2017

Nature

194

322

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Ca2+-selective transient receptor potential vanilloid subfamily member 6 (TRPV6) channels play a critical role in calcium uptake in epithelial tissues1–4. Altered TRPV6 expression is associated with a variety of human diseases5, including cancers6. TRPV6 channels are constitutively active1,7,8 and their open probability depends on the lipidic composition of the membrane, increasing significantly in the presence of phosphatidylinositol 4,5-bisphosphate (PIP2)7,9. We previously solved crystal structures of detergent-solubilized rat TRPV6 in the closed state10,11. Corroborating previous electrophysiological studies3, these structures demonstrated that the Ca2+ selectivity of TRPV6 arises from a ring of aspartate side chains in the selectivity filter that tightly binds Ca2+. However, it has remained unknown how TRPV6 channels open and close their pores for ion permeation. Here we present cryo-EM structures of human TRPV6 in the open and closed states. The channel selectivity filter adopts similar conformations in both states, consistent with its explicit role in ion permeation. The iris-like channel opening is accompanied by an α-to-π helical transition in the pore-lining S6 helices at an alanine hinge just below the selectivity filter. As a result of this transition, the S6 helices bend and rotate, exposing different residues to the ion channel pore in the open and closed states. This novel gating mechanism, which defines the constitutive activity of TRPV6, is unique for tetrameric ion channels and provides new structural insights for understanding their diverse roles in physiology and disease.

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Crystal Structure of a Mammalian Voltage-Dependent Shaker Family K ⁺ Channel

Cited by 2,053 publications

References 52 publications

Atomic structure of the eukaryotic intramembrane RAS methyltransferase ICMT

Atomic structure of the eukaryotic intramembrane RAS methyltransferase ICMT

The cytoplasmic coiled-coil mediates cooperative gating temperature sensitivity in the voltage-gated H+ channel Hv1

Opening of the human epithelial calcium channel TRPV6

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Crystal Structure of a Mammalian Voltage-Dependent Shaker Family K + Channel

Cited by 2,053 publications

References 52 publications

Atomic structure of the eukaryotic intramembrane RAS methyltransferase ICMT

Atomic structure of the eukaryotic intramembrane RAS methyltransferase ICMT

The cytoplasmic coiled-coil mediates cooperative gating temperature sensitivity in the voltage-gated H+ channel Hv1

Opening of the human epithelial calcium channel TRPV6

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Crystal Structure of a Mammalian Voltage-Dependent Shaker Family K ⁺ Channel