Chapter title |
Purification and Bicelle Crystallization for Structure Determination of the E. coli Outer Membrane Protein TamA
|
---|---|
Chapter number | 20 |
Book title |
The BAM Complex
|
Published in |
Methods in molecular biology, January 2015
|
DOI | 10.1007/978-1-4939-2871-2_20 |
Pubmed ID | |
Book ISBNs |
978-1-4939-2870-5, 978-1-4939-2871-2
|
Authors |
Fabian Gruss, Sebastian Hiller, Timm Maier, Gruss, Fabian, Hiller, Sebastian, Maier, Timm |
Abstract |
TamA is an Omp85 protein involved in autotransporter assembly in the outer membrane of Escherichia coli. It comprises a C-terminal 16-stranded transmembrane β-barrel as well as three periplasmic POTRA domains, and is a challenging target for structure determination. Here, we present a method for crystal structure determination of TamA, including recombinant expression in E. coli, detergent extraction, chromatographic purification, and bicelle crystallization in combination with seeding. As a result, crystals in space group P21212 are obtained, which diffract to 2.3 Å resolution. This protocol also serves as a template for structure determination of other outer membrane proteins, in particular of the Omp85 family. |
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