Structural basis of substrate recognition and translocation by human ABCA4

AbstractHuman ATP-binding cassette (ABC) subfamily A (ABCA) transporters mediate the transport of various lipid compounds across the membrane. Mutations in human ABCA transporters have been described to cause severe hereditary disorders associated with impaired lipid transport. However, little is known about the mechanistic details of substrate recognition and translocation by ABCA transporters. Here, we present three cryo-EM structures of human ABCA4, a retina-specific ABCA transporter, in distinct functional states at resolutions of 3.3–3.4 Å. In the nucleotide-free state, the two transmembrane domains (TMDs) exhibit a lateral-opening conformation, allowing the lateral entry of substrate from the lipid bilayer. The N-retinylidene-phosphatidylethanolamine (NRPE), the physiological lipid substrate of ABCA4, is sandwiched between the two TMDs in the luminal leaflet and is further stabilized by an extended loop from extracellular domain 1. In the ATP-bound state, the two TMDs display a closed conformation, which precludes the substrate binding. Our study provides a molecular basis to understand the mechanism of ABCA4-mediated NRPE recognition and translocation, and suggests a common ‘lateral access and extrusion’ mechanism for ABCA-mediated lipid transport.

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Structural basis of substrate recognition and translocation by human ABCA4

10.1101/2021.02.27.433154 ◽

2021 ◽

Author(s):

Tian Xie ◽

Zike Zhang ◽

Bowen Du ◽

Qi Fang ◽

Xin Gong

Keyword(s):

Substrate Recognition ◽

Bound State ◽

Lipid Transport ◽

Structural Basis ◽

Lateral Entry ◽

Closed Conformation ◽

Hereditary Disorders ◽

Extrusion Mechanism ◽

Functional States ◽

Lipid Compounds

AbstractHuman ATP-binding cassette (ABC) subfamily A (ABCA) transporters mediate the transport of various lipid compounds across the membrane. Mutations in human ABCA transporters have been described to cause severe hereditary disorders associated with impaired lipid transport. However, little is known about the mechanistic details of substrate recognition and translocation by ABCA transporters. Here, we present three cryo-EM structures of human ABCA4, a retinal-specific ABCA transporter, in distinct functional states at resolutions of 3.3-3.4 Å. In the nucleotide-free state, the two transmembrane domains (TMDs) exhibited a lateral-opening conformation, allowing the lateral entry of substrate from the lipid bilayer. N-retinylidene-phosphatidylethanolamine (NRPE), the physiological lipid substrate of ABCA4, is sandwiched between the two TMDs in the luminal leaflet and is further stabilized by an extended loop from extracellular domain 1. In the ATP-bound state, the two TMDs displayed an unprecedented closed conformation, which precludes the substrate binding. Our study provides a molecular basis to understand the mechanism of ABCA4-mediated NRPE recognition and translocation, and suggests a common ‘lateral access and extrusion’ mechanism for ABCA-mediated lipid transport.

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Structural basis of substrate recognition and translocation by human ABCD1

10.1101/2021.09.24.461565 ◽

2021 ◽

Author(s):

Zhipeng Chen ◽

Da Xu ◽

Liang Wang ◽

Cong-Zhao Zhou ◽

Wen-Tao Hou ◽

...

Keyword(s):

Genetic Disorder ◽

Substrate Recognition ◽

Acyl Chain ◽

Chain Fatty Acid ◽

Structural Basis ◽

Long Chain Fatty Acid ◽

Lateral Entry ◽

Binding Domains ◽

Acyl Chains ◽

Functional States

Human ATP-binding cassette (ABC) subfamily D transporter ABCD1 can transport CoA esters of saturated/monounsaturated long/very long chain fatty acid into the peroxisome for β-oxidation. Dysfunction of human ABCD1 causes X-linked adrenoleukodystrophy, which is a severe progressive genetic disorder affecting the nervous system. Nevertheless, the mechanistic details of substrate recognition and translocation by ABCD1 remains obscure. Here, we present three cryo-EM structures of human ABCD1 in distinct functional states. In the apo-form structure of 3.53 Å resolution, ABCD1 exhibits an inward-facing conformation, allowing the lateral entry of substrate from the lipid bilayer. In the 3.59 Å structure of substrate-bound ABCD1, two molecules of C22:0-CoA, the physiological substrate of ABCD1, is symmetrically bound in two transmembrane domains (TMDs). Each C22:0-CoA adopts a L-shape, with its CoA portion and acyl chain components bound to two TMDs respectively, resembling a pair of strings that pull the TMDs closer, resultantly generating a narrower outward-facing conformation. In the 2.79 Å ATP-bound ABCD1 structure, the two nucleotide-binding domains dimerize, leading to an outward-facing conformation, which opens the translocation cavity exit towards the peroxisome matrix side and releases the substrates. Our study provides a molecular basis to understand the mechanism of ABCD1-mediated substrate recognition and translocation, and suggests a unique binding pattern for amphipathic molecules with long acyl chains.

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Structural basis for the mechanisms of human presequence protease conformational switch and substrate recognition

10.21203/rs.3.rs-63850/v1 ◽

2020 ◽

Author(s):

Wenguang Liang ◽

Juwina wijaya ◽

Hui Wei ◽

Alex Noble ◽

Swansea Mo ◽

...

Keyword(s):

Citrate Synthase ◽

Substrate Binding ◽

Substrate Recognition ◽

Amyloid Β ◽

Body Motion ◽

Structural Basis ◽

Conformational Switch ◽

Closed Conformation ◽

Air Water Interface ◽

Open States

Abstract Presequence protease (PreP), a 117 kDa mitochondrial M16C metalloprotease vital for mitochondrial proteostasis, degrades presequence peptides cleaved off from nuclear-encoded proteins and other aggregation-prone peptides, such as amyloid β (Aβ). PreP structures have only been determined in a closed conformation; thus, the mechanisms of substrate binding and selectivity remain elusive. Here, we leverage advanced vitrification techniques to overcome the preferential denaturation of one of two ~55 kDa homologous domains of PreP caused by air-water interface adsorption, and thereby elucidate cryoEM structures of three apo-PreP open states along with Aβ- and citrate synthase presequence-bound PreP at 3.3 Å-4.6 Å resolution. Together with integrative biophysical and pharmacological approaches, these structures reveal the key stages of the PreP catalytic cycle and how the binding of substrates or PreP inhibitor drives the rigid body motion of the protein for substrate binding and catalysis. Together, our studies provide key mechanistic insights into M16C metalloproteases for future therapeutic innovations.

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Structural Basis and Sequence Rules for Substrate Recognition by Tankyrase Explain the Basis for Cherubism Disease

Cell ◽

10.1016/j.cell.2012.01.009 ◽

2012 ◽

Vol 148 (1-2) ◽

pp. 376

Author(s):

Sebastian Guettler ◽

Jose LaRose ◽

Evangelia Petsalaki ◽

Gerald Gish ◽

Andy Scotter ◽

...

Keyword(s):

Substrate Recognition ◽

Structural Basis ◽

Sequence Rules

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Structural basis of HMCES interactions with DNA reveals multivalent substrate recognition

10.1101/519595 ◽

2019 ◽

Author(s):

Levon Halabelian ◽

Mani Ravichandran ◽

Yanjun Li ◽

Hong Zheng ◽

L. Aravind ◽

...

Keyword(s):

Dna Damage ◽

Crystal Structures ◽

Genome Instability ◽

Substrate Recognition ◽

Catalytic Triad ◽

Structural Basis ◽

Replication Forks ◽

Abasic Sites ◽

Stalled Replication Forks ◽

Gapped Dna

ABSTRACTHMCES can covalently crosslink to abasic sites in single-stranded DNA at stalled replication forks to prevent genome instability. Here, we report crystal structures of the HMCES SRAP domain in complex with DNA-damage substrates, revealing interactions with both single-stranded and duplex segments of 3’ overhang DNA. HMCES may also bind gapped DNA and 5’ overhang structures to align single stranded abasic sites for crosslinking to the conserved Cys2 of its catalytic triad.

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Structural basis for osmotic regulation of the DNA binding properties of H-NS proteins

10.1101/757732 ◽

2019 ◽

Cited By ~ 2

Author(s):

Liang Qin ◽

Fredj Ben Bdira ◽

Yann G. J. Sterckx ◽

Alexander N. Volkov ◽

Jocelyne Vreede ◽

...

Keyword(s):

Dna Binding ◽

Electrostatic Interactions ◽

Intramolecular Interactions ◽

Structural Basis ◽

Binding Properties ◽

Family Protein ◽

Low Salt ◽

Functional States ◽

Mechanistic Basis ◽

Dna Binding Properties

AbstractH-NS proteins act as osmotic sensors translating changes in osmolarity into altered DNA binding properties, thus, regulating enterobacterial genome organization and genes transcription. The molecular mechanism underlying the switching process and its conservation among H-NS family members remains elusive.Here, we focus on the H-NS family protein MvaT from P. aeruginosa and demonstrate experimentally that its protomer exists in two different conformations, corresponding to two different functional states. In the half-opened state (dominant at low salt) the protein forms filaments along DNA, in the fully opened state (dominant at high salt) the protein bridges DNA. This switching is a direct effect of ionic strengths on electrostatic interactions between the appositively charged DNA binding and N-terminal domains of MvaT. The asymmetric charge distribution and intramolecular interactions are conserved among the H-NS family of proteins. Therefore, our study establishes a general paradigm for the molecular mechanistic basis of the osmosensitivity of H-NS proteins.

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