a-technique-for-stabilizing-membrane-proteins-in-nanodiscs

A Technique for Stabilizing Membrane Proteins in Nanodiscs

Many membrane proteins exhibit poor stability, loss of activity, and sample heterogeneity when extracted in detergent-based environments, posing ...

Fudan University

Carbonic anhydrase IV (Car4) is a newly identified receptor that allows adeno-associated virus (AAV) 9P31 to cross the blood-brain barrier and achieve efficient infection in the central nervous system (CNS) in mouse models. However, the molecular mechanism by which engineered AAV capsids with 7-mer insertion in the variable region (VR) VIII recognize these novel cellular receptors is unknown. Here we report the cryo-EM structures of AAV9P31 and its complex with Mus musculus Car4 at atomic resolution by utilizing the block-based reconstruction (BBR) method. The structures demonstrated that Car4 binds to the protrusions at 3-fold axes of the capsid. The inserted 7-mer extends into a hydrophobic region near the catalytic center of Car4 to form stable interactions. Mutagenesis studies also identified the key residues in Car4 responsible for the AAV9P31 interaction. These findings provide new insights into the novel receptor recognition mechanism of AAV generated by directed evolution and highlight the application of the BBR method to studying the virus-receptor molecular mechanism.

Structural basis of the recognition of adeno-associated virus by the neurological system-related receptor carbonic anhydrase IV.

Voltage-sensing phosphatases (VSPs) dephosphorylate phosphoinositide (PIP) signaling lipids in response to membrane depolarization. VSPs possess an S4-containing voltage sensor domain (VSD), resembling that of voltage-gated cation channels, and a lipid phosphatase domain (PD). The mechanism by which voltage turns on enzyme activity is unclear. Structural analysis and modeling suggest several sites of VSD-PD interaction that could couple voltage sensing to catalysis. Voltage clamp fluorometry reveals voltage-driven rearrangements in three sites implicated earlier in enzyme activation-the VSD-PD linker, gating loop and R loop-as well as the N-terminal domain, which has not yet been explored. N-terminus mutations perturb both rearrangements in the other segments and enzyme activity. Our results provide a model for a dynamic assembly by which S4 controls the catalytic site.

Coupling sensor to enzyme in the voltage sensing phosphatase.

Structure basis for sugar specificity of gustatory receptors in insects.

Pyrotinib, a novel pan-HER tyrosine kinase inhibitor, has demonstrated substantial anti-tumor activity in patients with NSCLC harboring HER2 mutations. Nevertheless, the inevitable resistance to pyrotinib necessitates an in-depth understanding of the underlying mechanisms.

AYVM to AYMM Transition on HER2 Exon 20 Insertion Induces Tyrosine Kinase Inhibitor Resistance in NSCLC.

Phosphorus in crucial for all living organisms. In vertebrate, cellular phosphate homeostasis is partly controlled by XPR1, a poorly characterized inositol pyrophosphate-dependent phosphate exporter. Here, we report the cryo-EM structure of human XPR1, which forms a loose dimer with 10 transmembrane helices (TM) in each protomer. The structure consists of a scaffold domain (TM1, TM3-4) and a core domain (TM2, TM5-10) structurally related to ion-translocating rhodopsins. Bound phosphate is observed in a tunnel within the core domain at a narrow point that separates the tunnel into intracellular and extracellular vestibules. This site contains a cluster of basic residues that coordinate phosphate and a conserved W573 essential for export function. Loss of inositol pyrophosphate binding is accompanied by structural movements in TM9 and the W573 sidechain, closing the extracellular vestibule and blocking phosphate export. These findings provide insight into XPR1 mechanism and pave the way for further in-depth XPR1 studies.

Structural basis of phosphate export by human XPR1.

TWIK-related halothane-inhibited potassium channel (THIK1) maintains the resting membrane potential and regulates potassium efflux in microglia. It is a potential therapeutic target for neurodegenerative disorders, neuropathic pain and inflammation. However, the mechanism underlying its function remains unclear. Here we used cryo-electron microscopy to solve the structures of full-length human THIK1, revealing two inner gates and a C-type selectivity filter gate, distinct from other two-pore-domain potassium channels. One inner gate, formed by a short helix in the distal C terminus, introduces a unique gating mechanism involving the distal cytoplasmic domain. The other, beneath the selectivity filter, is constricted by Y273 in the M4 helix, dividing the cavity. In addition, the selectivity filter gate is modulated by polyunsaturated fatty acids. These structural insights into THIK1 gating, through the distal C-terminal helices, hydrophilic residues and selectivity filter, advance our understanding of THIK1's role in microglial homeostasis and neuropathologies.

Article	Year
Structural basis of the recognition of adeno-associated virus by the neurological system-related receptor carbonic anhydrase IV. PLoS pathogens\| PubMed ID: 38315719	2024
Coupling sensor to enzyme in the voltage sensing phosphatase. Nature communications\| PubMed ID: 39080263	2024
Structure basis for sugar specificity of gustatory receptors in insects. Cell discovery\| PubMed ID: 39103325	2024
AYVM to AYMM Transition on HER2 Exon 20 Insertion Induces Tyrosine Kinase Inhibitor Resistance in NSCLC. Journal of thoracic oncology : official publication of the International Association for the Study of Lung Cancer\| PubMed ID: 39725168	2025
Structural basis of phosphate export by human XPR1. Nature communications\| PubMed ID: 39814721	2025
Gating mechanism of the two-pore-domain potassium channel THIK1. Nature structural & molecular biology\| PubMed ID: 40307591	2025

Baobin Li

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