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TitleSmall LEA proteins mitigate air-water interface damage to fragile cryo-EM samples during plunge freezing.
Journal, issue, pagesNat Commun, Vol. 15, Issue 1, Page 7705, Year 2024
Publish dateSep 4, 2024
AuthorsKaitlyn M Abe / Gan Li / Qixiang He / Timothy Grant / Ci Ji Lim /
PubMed AbstractAir-water interface (AWI) interactions during cryo-electron microscopy (cryo-EM) sample preparation cause significant sample loss, hindering structural biology research. Organisms like nematodes and ...Air-water interface (AWI) interactions during cryo-electron microscopy (cryo-EM) sample preparation cause significant sample loss, hindering structural biology research. Organisms like nematodes and tardigrades produce Late Embryogenesis Abundant (LEA) proteins to withstand desiccation stress. Here we show that these LEA proteins, when used as additives during plunge freezing, effectively mitigate AWI damage to fragile multi-subunit molecular samples. The resulting high-resolution cryo-EM maps are comparable to or better than those obtained using existing AWI damage mitigation methods. Cryogenic electron tomography reveals that particles are localized at specific interfaces, suggesting LEA proteins form a barrier at the AWI. This interaction may explain the observed sample-dependent preferred orientation of particles. LEA proteins offer a simple, cost-effective, and adaptable approach for cryo-EM structural biologists to overcome AWI-related sample damage, potentially revitalizing challenging projects and advancing the field of structural biology.
External linksNat Commun / PubMed:39231985 / PubMed Central
MethodsEM (single particle)
Resolution2.98 - 4.47 Å
Structure data

EMDB-43619: Human DNA polymerase alpha/primase - AavLEA1 (1:8 molar ratio)
Method: EM (single particle) / Resolution: 3.63 Å

EMDB-43620: Human PRC2 - AavLEA1 (1:40 molar ratio)
Method: EM (single particle) / Resolution: 3.82 Å

EMDB-43621: Human PRC2 - RvLEAM(short) (1:6 molar ratio)
Method: EM (single particle) / Resolution: 3.76 Å

EMDB-43622: Human PRC2 - RvLEAM(short) (1:6 molar ratio), 10 mM MgCl2
Method: EM (single particle) / Resolution: 4.18 Å

EMDB-43623: Human PRC2 - AavLEA1 (1:12 molar ratio)
Method: EM (single particle) / Resolution: 3.62 Å

43625

9c8u

EMDB-43625, PDB-9c8u:
Human PRC2 - RvLEAM (short) (1:6 molar ratio), cross-linked 10 min
Method: EM (single particle) / Resolution: 3.1 Å

EMDB-43626: Human DNA polymerase alpha/primase - RvLEAM(short) (1:6 molar ratio)
Method: EM (single particle) / Resolution: 4.47 Å

43627

9c8v

EMDB-43627: Human DNA polymerase alpha/primase - 4 mM CHAPSO
PDB-9c8v: Human DNA polymerase alpha/primase - CHAPSO (4 mM)
Method: EM (single particle) / Resolution: 3.39 Å

43628

8vy3

EMDB-43628, PDB-8vy3:
Human DNA polymerase alpha/primase - AavLEA1 (1:40 molar ratio)
Method: EM (single particle) / Resolution: 2.98 Å

EMDB-45273: Human PRC2 - cross-linked 10 min
Method: EM (single particle) / Resolution: 4.29 Å

Chemicals

ChemComp-ZN:
Unknown entry

ChemComp-SF4:
IRON/SULFUR CLUSTER

Source
  • homo sapiens (human)
KeywordsREPLICATION / Human DNA poly alpha/primase / AavLEA1 / air-water-interface / DNA BINDING PROTEIN / Human polycomb repressive complex 2 / RvLEAM / CHAPSO

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