title: Stoichiometric protein complex formation and over-expression using the prokaryotic native operon structure.
abstract: In prokaryotes, operon encoded proteins often form protein-protein complexes. Here, we show that the native structure of operons can be used to efficiently overexpress protein complexes. This study focuses on operons from mycobacteria and the use of Mycobacterium smegmatis as an expression host. We demonstrate robust and correct stoichiometric expression of dimers to higher oligomers. The expression efficacy was found to be largely independent of the intergenic distances. The strategy was successfully extended to express mycobacterial protein complexes in Escherichia coli, showing that the operon structure of gram-positive bacteria is also functional in gram-negative bacteria. The presented strategy could become a general tool for the expression of large quantities of pure prokaryotic protein complexes for biochemical and structural studies.
authors.affiliation: European Molecular Biology Laboratory (EMBL), Hamburg Outstation, c/o DESY, Hamburg, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Poulsen
authors.forename: Simon
authors.initials: S
authors.lastname: Holton
authors.forename: Arie
authors.initials: A
authors.lastname: Geerlof
authors.forename: Matthias
authors.initials: M
authors.lastname: Wilmanns
authors.forename: Young-Hwa
authors.initials: YH
authors.lastname: Song
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial; D006090:Gram-Negative Bacteria / Q000235:genetics; D006094:Gram-Positive Bacteria / Q000235:genetics; D008854:Microscopy, Electron; D008958:Models, Molecular; D046912:Multiprotein Complexes / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D009161:Mycobacterium / Q000235:genetics* / Q000378:metabolism; D009166:Mycobacterium leprae / Q000235:genetics / Q000378:metabolism; D020102:Mycobacterium smegmatis / Q000235:genetics / Q000378:metabolism; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000378:metabolism; D009876:Operon / Q000235:genetics*; D055503:Protein Multimerization
chemical_list: D001426:Bacterial Proteins; D046912:Multiprotein Complexes