Movie
Controller
Structure viewers
About Yorodumi Papers
+Search query
-Structure paper
| Title | An immobilized antibody-based affinity grid strategy for on-grid purification of target proteins enables high-resolution cryo-EM. |
|---|---|
| Journal, issue, pages | Commun Biol, Vol. 7, Issue 1, Page 715, Year 2024 |
| Publish date | Jun 10, 2024 |
 Authors | Qiaoyu Zhao / Xiaoyu Hong / Yanxing Wang / Shaoning Zhang / Zhanyu Ding / Xueming Meng / Qianqian Song / Qin Hong / Wanying Jiang / Xiangyi Shi / Tianxun Cai / Yao Cong /  |
| PubMed Abstract | In cryo-electron microscopy (cryo-EM), sample preparation poses a critical bottleneck, particularly for rare or fragile macromolecular assemblies and those suffering from denaturation and particle ...In cryo-electron microscopy (cryo-EM), sample preparation poses a critical bottleneck, particularly for rare or fragile macromolecular assemblies and those suffering from denaturation and particle orientation distribution issues related to air-water interface. In this study, we develop and characterize an immobilized antibody-based affinity grid (IAAG) strategy based on the high-affinity PA tag/NZ-1 antibody epitope tag system. We employ Pyr-NHS as a linker to immobilize NZ-1 Fab on the graphene oxide or carbon-covered grid surface. Our results demonstrate that the IAAG grid effectively enriches PA-tagged target proteins and overcomes preferred orientation issues. Furthermore, we demonstrate the utility of our IAAG strategy for on-grid purification of low-abundance target complexes from cell lysates, enabling atomic resolution cryo-EM. This approach greatly streamlines the purification process, reduces the need for large quantities of biological samples, and addresses common challenges encountered in cryo-EM sample preparation. Collectively, our IAAG strategy provides an efficient and robust means for combined sample purification and vitrification, feasible for high-resolution cryo-EM. This approach holds potential for broader applicability in both cryo-EM and cryo-electron tomography (cryo-ET). |
 External links | Commun Biol / PubMed:38858498 / PubMed Central |
| Methods | EM (single particle) |
| Resolution | 2.4 - 2.8 Å |
| Structure data |  EMDB-38142: Structure of CCT6-HR-ATP-AlFx  EMDB-38143: Structure of apoferritin  EMDB-38145: Consensus map of TBCA-apoferritin  EMDB-38147: Structure of CCT6-HR  EMDB-39651: Structure of the focused refined TBCA-apoferritin |
| Source |
|
Saccharomyces cerevisiae (brewer's yeast)
Homo sapiens (human)