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Purification, refolding and characterization of the trimeric Omp2a outer membrane porin from Brucella melitensis.
- Source :
-
Protein expression and purification [Protein Expr Purif] 2012 Jun; Vol. 83 (2), pp. 198-204. Date of Electronic Publication: 2012 Apr 17. - Publication Year :
- 2012
-
Abstract
- Brucella melitensis is a gram-negative bacteria known to cause brucellosis and to produce severe infections in humans. Whilst brucella's outer membrane proteins have been extensively studied due to their potential role as antigens or virulence factors, their function is still poorly understood at the structural level, as the 3D structure of Brucella β-barrel membrane proteins are still unknown. In this context, the B. melitensis trimeric Omp2a porin has been overexpressed and refolded in n-dodecyl-β-d-maltopyranoside. We here show that this refolding process is insensitive to urea but is temperature- and ionic strength-dependent. Reassembled species were characterized by fluorescence, size-exclusion chromatography and circular dichroism. A refolding mechanism is proposed, suggesting that Omp2a first refolds under a monomeric form and then self-associates into a trimeric state. This first complete in vitro refolding of a membrane protein from B. melitensis shall eventually lead to functional and 3D structure determination.<br /> (Copyright © 2012 Elsevier Inc. All rights reserved.)
- Subjects :
- Amino Acid Sequence
Bacterial Proteins genetics
Bacterial Proteins metabolism
Base Sequence
Brucella melitensis chemistry
Brucella melitensis metabolism
Circular Dichroism
Escherichia coli genetics
Maltose analogs & derivatives
Maltose chemistry
Molecular Sequence Data
Porins genetics
Porins metabolism
Protein Refolding
Recombinant Proteins genetics
Recombinant Proteins metabolism
Temperature
Bacterial Proteins chemistry
Brucella melitensis genetics
Porins chemistry
Recombinant Proteins chemistry
Subjects
Details
- Language :
- English
- ISSN :
- 1096-0279
- Volume :
- 83
- Issue :
- 2
- Database :
- MEDLINE
- Journal :
- Protein expression and purification
- Publication Type :
- Academic Journal
- Accession number :
- 22538317
- Full Text :
- https://doi.org/10.1016/j.pep.2012.04.003