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Long shelf-life streptavidin support-films suitable for electron microscopy of biological macromolecules.

ABSTRACT: We describe a rapid and convenient method of growing streptavidin (SA) monolayer crystals directly on holey-carbon EM grids. As expected, these SA monolayer crystals retain their biotin-binding function and crystalline order through a cycle of embedding in trehalose and, later, its removal. This fact allows one to prepare, and store for later use, EM grids on which SA monolayer crystals serve as an affinity substrate for preparing specimens of biological macromolecules. In addition, we report that coating the lipid-tail side of trehalose-embedded monolayer crystals with evaporated carbon appears to improve the consistency with which well-ordered, single crystals are observed to span over entire, 2?m holes of the support films. Randomly biotinylated 70S ribosomes are used as a test specimen to show that these support films can be used to obtain a high-resolution cryo-EM structure.

SUBMITTER: Han BG

PROVIDER: S-EPMC4943657 | biostudies-literature | 2016 Aug

REPOSITORIES: biostudies-literature

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Long shelf-life streptavidin support-films suitable for electron microscopy of biological macromolecules.

Han Bong-Gyoon BG Watson Zoe Z Kang Hannah H Pulk Arto A Downing Kenneth H KH Cate Jamie J Glaeser Robert M RM

Journal of structural biology 20160615 2

We describe a rapid and convenient method of growing streptavidin (SA) monolayer crystals directly on holey-carbon EM grids. As expected, these SA monolayer crystals retain their biotin-binding function and crystalline order through a cycle of embedding in trehalose and, later, its removal. This fact allows one to prepare, and store for later use, EM grids on which SA monolayer crystals serve as an affinity substrate for preparing specimens of biological macromolecules. In addition, we report th ...[more]

PMID: 27320699

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