Expression of membrane proteins from Mycobacterium tuberculosis in Escherichia coli as fusions with maltose binding protein

• Published in: Protein expression and purification 2007-05, Vol.53 (1), p.24-30
• Main Authors: Korepanova, A., Moore, J.D., Nguyen, H.B., Hua, Y., Cross, T.A., Gao, F.
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Bacterial Proteins - chemistry; Bacterial Proteins - genetics; Bacterial Proteins - isolation & purification; Bacterial Proteins - metabolism; Carrier Proteins - metabolism; Cloning, Molecular; Escherichia coli; Escherichia coli - genetics; Escherichia coli - metabolism; Fusion proteins; Gene Expression; Histidine - chemistry; Inclusion Bodies - metabolism; Maltose binding protein; Maltose-Binding Proteins; Membrane protein expression; Membrane Proteins - chemistry; Membrane Proteins - genetics; Membrane Proteins - isolation & purification; Membrane Proteins - metabolism; Molecular Sequence Data; Molecular Weight; Mycobacterium tuberculosis; Mycobacterium tuberculosis - chemistry; Plasmids; Polymerase Chain Reaction; Protein Conformation; Recombinant Fusion Proteins - chemistry; Recombinant Fusion Proteins - isolation & purification; Recombinant Fusion Proteins - metabolism; Solubility
• ISSN: 1046-5928; 1096-0279
• DOI: 10.1016/j.pep.2006.11.022

Sixteen of 22 low molecular weight integral membrane proteins from Mycobacterium tuberculosis with previously poor or undetectable levels of expression were expressed in Escherichia coli as fusions with both the maltose binding protein (MBP) and a His 8-tag. Sixty-eight percent of targeted proteins were expressed in high yield (>30 mg/L) in soluble and/or inclusion body form. Thrombin cleavage of the MBP fusion protein was successful for 10 of 13 proteins expressed as soluble proteins and for three proteins expressed only as inclusion bodies. The use of autoinduction growth media increased yields over Luria-Bertani (LB) growth media in 75% of the expressed proteins. Expressing integral membrane proteins with yields suitable for structural studies from a set of previously low and non-expressing proteins proved highly successful upon attachment of the maltose binding protein as a fusion tag.