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TitleExploring high-resolution cryo-ET and subtomogram averaging capabilities of contemporary DEDs.
Journal, issue, pagesJ Struct Biol, Vol. 214, Issue 2, Page 107852, Year 2022
Publish dateMar 26, 2022
AuthorsMartin Obr / Wim J H Hagen / Robert A Dick / Lingbo Yu / Abhay Kotecha / Florian K M Schur /
PubMed AbstractThe potential of energy filtering and direct electron detection for cryo-electron microscopy (cryo-EM) has been well documented. Here, we assess the performance of recently introduced hardware for ...The potential of energy filtering and direct electron detection for cryo-electron microscopy (cryo-EM) has been well documented. Here, we assess the performance of recently introduced hardware for cryo-electron tomography (cryo-ET) and subtomogram averaging (STA), an increasingly popular structural determination method for complex 3D specimens. We acquired cryo-ET datasets of EIAV virus-like particles (VLPs) on two contemporary cryo-EM systems equipped with different energy filters and direct electron detectors (DED), specifically a Krios G4, equipped with a cold field emission gun (CFEG), Thermo Fisher Scientific Selectris X energy filter, and a Falcon 4 DED; and a Krios G3i, with a Schottky field emission gun (XFEG), a Gatan Bioquantum energy filter, and a K3 DED. We performed constrained cross-correlation-based STA on equally sized datasets acquired on the respective systems. The resulting EIAV CA hexamer reconstructions show that both systems perform comparably in the 4-6 Å resolution range based on Fourier-Shell correlation (FSC). In addition, by employing a recently introduced multiparticle refinement approach, we obtained a reconstruction of the EIAV CA hexamer at 2.9 Å. Our results demonstrate the potential of the new generation of energy filters and DEDs for STA, and the effects of using different processing pipelines on their STA outcomes.
External linksJ Struct Biol / PubMed:35351542
MethodsEM (subtomogram averaging) / EM (tomography)
Resolution2.9 - 3.6 Å
Structure data

EMDB-14031:
Structure of the EIAV CA-SP hexamer (C6), acquired on a Krios 4, Selectris/Falcon4, processed with Relion/M
Method: EM (subtomogram averaging) / Resolution: 2.9 Å

EMDB-14032: Structure of the EIAV CA-SP2 hexamer (C2), acquired on Krios G4, Selectris/Falcon4, processed with Relion/M
Method: EM (subtomogram averaging) / Resolution: 3.2 Å

EMDB-14033:
Structure of the EIAV CA-SP hexamer (C6), acquired on Krios 3Gi/Bioquantum K3, processed with Relion/M
Method: EM (subtomogram averaging) / Resolution: 3.3 Å

EMDB-14034: Structure of the EIAV CA-SP hexamer (C2), acquired on Krios 3Gi/Gatan Bioquantum K3, processed with Relion/M
Method: EM (subtomogram averaging) / Resolution: 3.6 Å

EMDB-14035:
Structure of the EIAV CA-SP hexamer (C6), acquired on Krios4, Selectris/Falcon4, processed with AV3
Method: EM (subtomogram averaging) / Resolution: 3.4 Å

EMDB-14036:
Cryo-electron tomogram of EIAV CASPNC VLPs, acquired on Krios G4, Selectris/Falcon4
Method: EM (tomography)

EMDB-14037:
Cryo-electron tomogram of EIAV CASPNC VLPs, acquired on Krios 3Gi, Bioquantum K3
Method: EM (tomography)

Source
  • Equine infectious anemia virus
  • Escherichia coli (E. coli)

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