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Title: 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 μm 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

Authors:
 [1];  [2];  [1];  [3];  [1];  [4];  [1]
  1. Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Molecular Biophysics and Integrated Bioimaging Division
  2. Univ. of California, Berkeley, CA (United States). Dept. of Chemistry
  3. Univ. of California, Berkeley, CA (United States). Dept. of Molecular and Cell Biology and California Inst. of Quantitative Biosciense
  4. Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Molecular Biophysics and Integrated Bioimaging Division; Univ. of California, Berkeley, CA (United States). Dept. of Chemistry; Univ. of California, Berkeley, CA (United States). Dept. of Molecular and Cell Biology and California Inst. of Quantitative Biosciense
Publication Date:
Research Org.:
Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)
Sponsoring Org.:
USDOE; National Institutes of Health (NIH)
OSTI Identifier:
1378360
Grant/Contract Number:  
AC02-05CH11231; R01-GM083039; P01-GM051487; R01-GM065050
Resource Type:
Journal Article: Accepted Manuscript
Journal Name:
Journal of Structural Biology
Additional Journal Information:
Journal Volume: 195; Journal Issue: 2; Journal ID: ISSN 1047-8477
Publisher:
Elseiver
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES

Citation Formats

Han, Bong-Gyoon, Watson, Zoe, Kang, Hannah, Pulk, Arto, Downing, Kenneth H., Cate, Jamie, and Glaeser, Robert M.. Long shelf-life streptavidin support-films suitable for electron microscopy of biological macromolecules. United States: N. p., 2016. Web. doi:10.1016/j.jsb.2016.06.009.
Han, Bong-Gyoon, Watson, Zoe, Kang, Hannah, Pulk, Arto, Downing, Kenneth H., Cate, Jamie, & Glaeser, Robert M.. Long shelf-life streptavidin support-films suitable for electron microscopy of biological macromolecules. United States. doi:10.1016/j.jsb.2016.06.009.
Han, Bong-Gyoon, Watson, Zoe, Kang, Hannah, Pulk, Arto, Downing, Kenneth H., Cate, Jamie, and Glaeser, Robert M.. Wed . "Long shelf-life streptavidin support-films suitable for electron microscopy of biological macromolecules". United States. doi:10.1016/j.jsb.2016.06.009. https://www.osti.gov/servlets/purl/1378360.
@article{osti_1378360,
title = {Long shelf-life streptavidin support-films suitable for electron microscopy of biological macromolecules},
author = {Han, Bong-Gyoon and Watson, Zoe and Kang, Hannah and Pulk, Arto and Downing, Kenneth H. and Cate, Jamie and Glaeser, Robert M.},
abstractNote = {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 μm 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},
doi = {10.1016/j.jsb.2016.06.009},
journal = {Journal of Structural Biology},
number = 2,
volume = 195,
place = {United States},
year = {Wed Jun 15 00:00:00 EDT 2016},
month = {Wed Jun 15 00:00:00 EDT 2016}
}

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Cited by: 2 works
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