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  LptC from Anabaena sp. PCC 7120: Expression, purification and crystallization

Ngo, G., Centola, M., Krasnoselska, G., Pogoryelov, D., Yildiz, Ö., & Schleiff, E. (2020). LptC from Anabaena sp. PCC 7120: Expression, purification and crystallization. Protein Expression and Purification, 175: 105689. doi:10.1016/j.pep.2020.105689.

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 Creators:
Ngo, Giang1, Author
Centola, Martin2, Author           
Krasnoselska, Ganna3, Author
Pogoryelov, Denys2, Author           
Yildiz, Özkan2, Author                 
Schleiff, Enrico1, 4, 5, Author
Affiliations:
1Institute for Molecular Biosciences, Goethe University Frankfurt, 60438 Frankfurt, Germany, ou_persistent22              
2Department of Structural Biology, Max Planck Institute of Biophysics, Max Planck Society, ou_2068291              
3Institute for Biochemistry, Goethe University Frankfurt, 60438 Frankfurt, Germany, ou_persistent22              
4Cluster of Excellence Macromolecular Complexes, 60438 Frankfurt, Germany, ou_persistent22              
5Frankfurt Institute for Advanced Studies, 60438 Frankfurt, Germany, ou_persistent22              

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Free keywords: Lipid A; lipopolysaccharides; periplasmic transfer; protein purification; crystal diffraction; cyanobacteria
 Abstract: Lipopolysaccharides are central elements of the outer leaflet of the outer membrane of Gram-negative bacteria and as such, of cyanobacteria. In the past, the structural analysis of the system in proteobacteria like Escherichia coli has contributed to a deep understanding of the transport of lipopolysaccharides from plasma membrane to the outer membrane. While many components of the transport system are conserved between proteobacteria and cyanobacteria, the periplasmic LptC appears to be distinct. The cyanobacterial proteins are twice as long as the proteobacterial proteins or proteins from firmicutes. This prompted the question whether the structure of the cyanobacterial proteins is comparable the one of the proteobacterial proteins. To address this question, we expressed LptC from Anabaena sp. PCC 7120 in E. coli as truncated protein without the transmembrane segment. We purified the protein utilizing HIS-tag based affinity chromatography and polished the protein after removal of the tag by size exclusion chromatography. The purified recombinant protein was crystallized by the sitting-drop vapor diffusion technique and best crystals, despite being twinned, diffracted to a resolution of 2.6 Å.

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Language(s): eng - English
 Dates: 2020-06-052020-03-172020-06-162020-07-192020-11
 Publication Status: Issued
 Pages: 5
 Publishing info: -
 Table of Contents: -
 Rev. Type: Peer
 Identifiers: DOI: 10.1016/j.pep.2020.105689
 Degree: -

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Title: Protein Expression and Purification
Source Genre: Journal
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Publ. Info: San Diego : Academic Press
Pages: - Volume / Issue: 175 Sequence Number: 105689 Start / End Page: - Identifier: ISSN: 1046-5928
CoNE: https://pure.mpg.de/cone/journals/resource/954922650158