Cryo-EM structures of human Cx36/GJD2 neuronal gap junction channel.

Connexin 36 (Cx36) is responsible for signal transmission in electrical synapses by forming interneuronal gap junctions. Despite the critical role of Cx36 in normal brain function, the molecular architecture of the Cx36 gap junction channel (GJC) is unknown. Here, we determine cryo-electron microscopy structures of Cx36 GJC at 2.

Conformational changes in the human Cx43/GJA1 gap junction channel visualized using cryo-EM.

Connexin family proteins assemble into hexameric hemichannels in the cell membrane. The hemichannels dock together between two adjacent membranes to form gap junction intercellular channels (GJIChs). We report the cryo-electron microscopy structures of Cx43 GJICh, revealing the dynamic equilibrium state of various channel conformations in detergents and lipid nanodiscs.

Heterologous Expression and Purification of a CRISPR-Cas9-Based Adenine Base Editor.

CRISPR-cas9-guided adenine base editors (ABEs) site-specifically convert the A-T base pair to G-C base pair in genomic DNA. The intracellular delivery of ABE proteins preassembled with guide RNAs (gRNAs) has shown greatly reduced off-target effects compared with that of plasmids or viral vectors containing ABE and gRNA-encoding sequences. For efficient gene editing by the ribonucleoprotein delivery method, the ABE-gRNA complexes need to be prepared in high purity and quantity.

High-purity production and precise editing of DNA base editing ribonucleoproteins.

Ribonucleoprotein (RNP) complex-mediated base editing is expected to be greatly beneficial because of its reduced off-target effects compared to plasmid- or viral vector-mediated gene editing, especially in therapeutic applications. However, production of recombinant cytosine base editors (CBEs) or adenine base editors (ABEs) with ample yield and high purity in bacterial systems is challenging. Here, we obtained highly purified CBE/ABE proteins from a human cell expression system and showed that CBE/ABE RNPs exhibited different editing patterns (i.

Application of Solution NMR to Structural Studies on α-Helical Integral Membrane Proteins.

A large portion of proteins in living organisms are membrane proteins which play critical roles in the biology of the cell, from maintenance of the biological membrane integrity to communication of cells with their surroundings. To understand their mechanism of action, structural information is essential. Nevertheless, structure determination of transmembrane proteins is still a challenging area, even though recently the number of deposited structures of membrane proteins in the PDB has rapidly increased thanks to the efforts using X-ray crystallography, electron microscopy, and solid and solution nuclear magnetic resonance (NMR) technology.