Improved mycobacterial protein production using a Mycobacterium smegmatis groEL1ؤC expression strain
- نوع فایل : کتاب
- زبان : انگلیسی
- مؤلف : Elke E Noens1*, Chris Williams1, Madhankumar Anandhakrishnan1, Christian Poulsen2, Matthias T Ehebauer1 and Matthias Wilmanns1
- چاپ و سال / کشور: 2011
Description
Background: The non-pathogenic bacterium Mycobacterium smegmatis is widely used as a near-native expression host for the purification of Mycobacterium tuberculosis proteins. Unfortunately, the Hsp60 chaperone GroEL1, which is relatively highly expressed, is often co-purified with polyhistidine-tagged recombinant proteins as a major contaminant when using this expression system. This is likely due to a histidine-rich C-terminus in GroEL1. Results: In order to improve purification efficiency and yield of polyhistidine-tagged mycobacterial target proteins, we created a mutant version of GroEL1 by removing the coding sequence for the histidine-rich C-terminus, termed GroEL1ؤC. GroEL1ؤC, which is a functional protein, is no longer able to bind nickel affinity beads. Using a selection of challenging test proteins, we show that GroEL1ؤC is no longer present in protein samples purified from the groEL1ؤC expression strain and demonstrate the feasibility and advantages of purifying and characterising proteins produced using this strain. Conclusions: This novel Mycobacterium smegmatis expression strain allows efficient expression and purification of mycobacterial proteins while concomitantly removing the troublesome contaminant GroEL1 and consequently increasing the speed and efficiency of protein purification.
Noens et al. BMC Biotechnology 2011, 11:27 http://www.biomedcentral.com/1472-6750/11/27
