Bing Rao

All Publications

• The cryo-EM structure of the human ERAD retrotranslocation complex. Science advances Rao, B., Wang, Q., Yao, D., Xia, Y., Li, W., Xie, Y., Li, S., Cao, M., Shen, Y., Qin, A., Zhao, J., Cao, Y. 2023; 9 (41): eadi5656

  Abstract

  Endoplasmic reticulum-associated degradation (ERAD) maintains protein homeostasis by retrieving misfolded proteins from the endoplasmic reticulum (ER) lumen into the cytosol for degradation. The retrotranslocation of misfolded proteins across the ER membrane is an energy-consuming process, with the detailed transportation mechanism still needing clarification. We determined the cryo-EM structures of the hetero-decameric complex formed by the Derlin-1 tetramer and the p97 hexamer. It showed an intriguing asymmetric complex and a putative coordinated squeezing movement in Derlin-1 and p97 parts. With the conformational changes of p97 induced by its ATP hydrolysis activities, the Derlin-1 channel could be torn into a "U" shape with a large opening to the lipidic environment, thereby forming an entry for the substrates in the ER membrane. The EM analysis showed that p97 formed a functional protein complex with Derlin-1, revealing the coupling mechanism between the ERAD retrotranslocation and the ATP hydrolysis activities.

  View details for DOI 10.1126/sciadv.adi5656

  View details for PubMedID 37831771

  View details for PubMedCentralID PMC10575581

• The structural pathology for hypophosphatasia caused by malfunctional tissue non-specific alkaline phosphatase NATURE COMMUNICATIONS Yu, Y., Rong, K., Yao, D., Zhang, Q., Cao, X., Rao, B., Xia, Y., Lu, Y., Shen, Y., Yao, Y., Xu, H., Ma, P., Cao, Y., Qin, A. 2023; 14 (1): 4048

  Abstract

  Hypophosphatasia (HPP) is a metabolic bone disease that manifests as developmental abnormalities in bone and dental tissues. HPP patients exhibit hypo-mineralization and osteopenia due to the deficiency or malfunction of tissue non-specific alkaline phosphatase (TNAP), which catalyzes the hydrolysis of phosphate-containing molecules outside the cells, promoting the deposition of hydroxyapatite in the extracellular matrix. Despite the identification of hundreds of pathogenic TNAP mutations, the detailed molecular pathology of HPP remains unclear. Here, to address this issue, we determine the crystal structures of human TNAP at near-atomic resolution and map the major pathogenic mutations onto the structure. Our study reveals an unexpected octameric architecture for TNAP, which is generated by the tetramerization of dimeric TNAPs, potentially stabilizing the TNAPs in the extracellular environments. Moreover, we use cryo-electron microscopy to demonstrate that the TNAP agonist antibody (JTALP001) forms a stable complex with TNAP by binding to the octameric interface. The administration of JTALP001 enhances osteoblast mineralization and promoted recombinant TNAP-rescued mineralization in TNAP knockout osteoblasts. Our findings elucidate the structural pathology of HPP and highlight the therapeutic potential of the TNAP agonist antibody for osteoblast-associated bone disorders.

  View details for DOI 10.1038/s41467-023-39833-3

  View details for Web of Science ID 001025242000014

  View details for PubMedID 37422472

  View details for PubMedCentralID PMC10329691

• The structural basis of the pH-homeostasis mediated by the Cl-/HCO3- exchanger, AE2 NATURE COMMUNICATIONS Zhang, Q., Jian, L., Yao, D., Rao, B., Xia, Y., Hu, K., Li, S., Shen, Y., Cao, M., Qin, A., Zhao, J., Cao, Y. 2023; 14 (1): 1812

  Abstract

  The cell maintains its intracellular pH in a narrow physiological range and disrupting the pH-homeostasis could cause dysfunctional metabolic states. Anion exchanger 2 (AE2) works at high cellular pH to catalyze the exchange between the intracellular HCO3- and extracellular Cl-, thereby maintaining the pH-homeostasis. Here, we determine the cryo-EM structures of human AE2 in five major operating states and one transitional hybrid state. Among those states, the AE2 shows the inward-facing, outward-facing, and intermediate conformations, as well as the substrate-binding pockets at two sides of the cell membrane. Furthermore, critical structural features were identified showing an interlock mechanism for interactions among the cytoplasmic N-terminal domain and the transmembrane domain and the self-inhibitory effect of the C-terminal loop. The structural and cell-based functional assay collectively demonstrate the dynamic process of the anion exchange across membranes and provide the structural basis for the pH-sensitive pH-rebalancing activity of AE2.

  View details for DOI 10.1038/s41467-023-37557-y

  View details for Web of Science ID 000980769900014

  View details for PubMedID 37002221

  View details for PubMedCentralID PMC10066210

• C-terminal deletion-induced condensation sequesters AID from IgH targets in immunodeficiency EMBO JOURNAL Xie, X., Gan, T., Rao, B., Zhang, W., Panchakshari, R. A., Yang, D., Ji, X., Cao, Y., Alt, F. W., Meng, F., Hu, J. 2022; 41 (11): e109324

  Abstract

  In activated B cells, activation-induced cytidine deaminase (AID) generates programmed DNA lesions required for antibody class switch recombination (CSR), which may also threaten genome integrity. AID dynamically shuttles between cytoplasm and nucleus, and the majority stays in the cytoplasm due to active nuclear export mediated by its C-terminal peptide. In immunodeficient-patient cells expressing mutant AID lacking its C-terminus, a catalytically active AID-delC protein accumulates in the nucleus but nevertheless fails to support CSR. To resolve this apparent paradox, we dissected the function of AID-delC proteins in the CSR process and found that they cannot efficiently target antibody genes. We demonstrate that AID-delC proteins form condensates both in vivo and in vitro, dependent on its N-terminus and on a surface arginine-rich patch. Co-expression of AID-delC and wild-type AID leads to an unbalanced nuclear AID-delC/AID ratio, with AID-delC proteins able to trap wild-type AID in condensates, resulting in a dominant-negative phenotype that could contribute to immunodeficiency. The co-condensation model of mutant and wild-type proteins could be an alternative explanation for the dominant-negative effect in genetic disorders.

  View details for DOI 10.15252/embj.2021109324

  View details for Web of Science ID 000787322200001

  View details for PubMedID 35471583

  View details for PubMedCentralID PMC9156971

• The structural basis for the phospholipid remodeling by lysophosphatidylcholine acyltransferase 3 NATURE COMMUNICATIONS Zhang, Q., Yao, D., Rao, B., Jian, L., Chen, Y., Hu, K., Xia, Y., Li, S., Shen, Y., Qin, A., Zhao, J., Zhou, L., Lei, M., Jiang, X., Cao, Y. 2021; 12 (1): 6869

  Abstract

  As the major component of cell membranes, phosphatidylcholine (PC) is synthesized de novo in the Kennedy pathway and then undergoes extensive deacylation-reacylation remodeling via Lands' cycle. The re-acylation is catalyzed by lysophosphatidylcholine acyltransferase (LPCAT) and among the four LPCAT members in human, the LPCAT3 preferentially introduces polyunsaturated acyl onto the sn-2 position of lysophosphatidylcholine, thereby modulating the membrane fluidity and membrane protein functions therein. Combining the x-ray crystallography and the cryo-electron microscopy, we determined the structures of LPCAT3 in apo-, acyl donor-bound, and acyl receptor-bound states. A reaction chamber was revealed in the LPCAT3 structure where the lysophosphatidylcholine and arachidonoyl-CoA were positioned in two tunnels connected near to the catalytic center. A side pocket was found expanding the tunnel for the arachidonoyl CoA and holding the main body of arachidonoyl. The structural and functional analysis provides the basis for the re-acylation of lysophosphatidylcholine and the substrate preference during the reactions.

  View details for DOI 10.1038/s41467-021-27244-1

  View details for Web of Science ID 000722866700057

  View details for PubMedID 34824256

  View details for PubMedCentralID PMC8617236

• The cryo-EM structure of an ERAD protein channel formed by tetrameric human Derlin-1 SCIENCE ADVANCES Rao, B., Li, S., Yao, D., Wang, Q., Xia, Y., Jia, Y., Shen, Y., Cao, Y. 2021; 7 (10)

  Abstract

  Endoplasmic reticulum-associated degradation (ERAD) is a process directing misfolded proteins from the ER lumen and membrane to the degradation machinery in the cytosol. A key step in ERAD is the translocation of ER proteins to the cytosol. Derlins are essential for protein translocation in ERAD, but the mechanism remains unclear. Here, we solved the structure of human Derlin-1 by cryo-electron microscopy. The structure shows that Derlin-1 forms a homotetramer that encircles a large tunnel traversing the ER membrane. The tunnel has a diameter of about 12 to 15 angstroms, large enough to allow an α helix to pass through. The structure also shows a lateral gate within the membrane, providing access of transmembrane proteins to the tunnel, and thus, human Derlin-1 forms a protein channel for translocation of misfolded proteins. Our structure is different from the monomeric yeast Derlin structure previously reported, which forms a semichannel with another protein.

  View details for DOI 10.1126/sciadv.abe8591

  View details for Web of Science ID 000625411900012

  View details for PubMedID 33658201

  View details for PubMedCentralID PMC7929502