The recent outbreaks of SARS-CoV-2 pose a global health emergency. The SARS-CoV-2 trimeric spike (S) glycoprotein interacts with the human ACE2 receptor to mediate viral entry into host cells. We report the cryo-EM structures of a tightly closed SARS-CoV-2 S trimer with packed fusion peptide and an ACE2-bound S trimer at 2.7- and 3.8-Å resolution, respectively. Accompanying ACE2 binding to the up receptor-binding domain (RBD), the associated ACE2-RBD exhibits continuous swing motions. Notably, the SARS-CoV-2 S trimer appears much more sensitive to the ACE2 receptor than the SARS-CoV S trimer regarding receptor-triggered transformation from the closed prefusion state to the fusion-prone open state, potentially contributing to the superior infectivity of SARS-CoV-2. We defined the RBD T470-T478 loop and Y505 as viral determinants for specific recognition of SARS-CoV-2 RBD by ACE2. Our findings depict the mechanism of ACE2-induced S trimer conformational transitions from the ground prefusion state toward the postfusion state, facilitating development of anti–SARS-CoV-2 vaccines and therapeutics.

Mitochondria from multiple, eukaryotic clades uptake and buffer large amounts of calcium (Ca2+) via an inner membrane transporter called the uniporter. Early studies demonstrated that this transport requires a mitochondrial membrane potential and that the uniporter is itself Ca2+ activated, and blocked by ruthenium red or Ru3601. Later, electrophysiological studies demonstrated that the uniporter is an ion channel with remarkably high conductance and selectivity2. Ca2+ entry into mitochondria is also known to activate the TCA cycle and appears to be critical for matching ATP production in mitochondria with its cytosolic demand3. MCU (mitochondrial calcium uniporter) is the pore forming and Ca2+ conducting subunit of the uniporter, but its primary sequence does not resemble any calcium channel known to date. Here, we report the structure of the core region of MCU, determined using nuclear magnetic resonance (NMR) and electron microscopy (EM). MCU is a homo-oligomer with the second transmembrane helix forming a hydrophilic pore across the membrane. The channel assembly represents a new solution of ion channel architecture and is stabilized by a coiled coil motif protruding in the mitochondrial matrix. The critical DxxE motif forms the pore entrance featuring two carboxylate rings, which appear to be the selectivity filter based on the ring dimensions and functional mutagenesis. To our knowledge, this is one of the largest structures characterized by NMR, which provides a structural blueprint for understanding the function of this channel.

CRISPR-Cas9 technology has been widely used for genome engineering. Its RNA-guided endonuclease Cas9 binds specifically to target DNA and then cleaves the two DNA strands with HNH and RuvC nuclease domains. However, structural information regarding the DNA cleavage-activating state of two nuclease domains remains sparse. Here, we report a 5.2 Å cryo-EM structure of Cas9 in complex with sgRNA and target DNA. This structure reveals a conformational state of Cas9 in which the HNH domain is closest to the DNA cleavage site. Compared with two known HNH states, our structure shows that the HNH active site moves toward the cleavage site by about 25 and 13 Å, respectively. In combination with EM-based molecular dynamics simulations, we show that residues of the nuclease domains in our structure could form cleavage-compatible conformations with the target DNA. Together, these results strongly suggest that our cryo-EM structure resembles a DNA cleavage-activating architecture of Cas9.

The eukaryotic chaperonin TRiC (or CCT) assists in the folding of 10% of cytosolic proteins. Here we present two cryo-EM structures of Saccharomyces cerevisiae TRiC in a newly identified nucleotide partially preloaded (NPP) state and in the ATP-bound state, at 4.7-Å and 4.6-Å resolution, respectively. Through inner-subunit eGFP tagging, we identified the subunit locations in open-state TRiC and found that the CCT2 subunit pair forms an unexpected Z shape. ATP binding induces a dramatic conformational change on the CCT2 side, thereby suggesting that CCT2 plays an essential role in TRiC allosteric cooperativity. Our structural and biochemical data reveal a staggered ATP binding mechanism of TRiC with preloaded nucleotide on the CCT6 side of NPP-TRiC and demonstrate that TRiC has evolved into a complex that is structurally divided into two sides. This work offers insight into how the TRiC nucleotide cycle coordinates with its mechanical cycle in preparing folding intermediates for further productive folding.