Improving membrane protein expression and function using genomic edits.
Sci Rep
; 7(1): 13030, 2017 10 12.
Article
in En
| MEDLINE
| ID: mdl-29026162
ABSTRACT
Expression of membrane proteins often leads to growth inhibition and perturbs central metabolism and this burden varies with the protein being overexpressed. There are also known strain backgrounds that allow greater expression of membrane proteins but that differ in efficacy across proteins. We hypothesized that for any membrane protein, it may be possible to identify a modified strain background where its expression can be accommodated with less burden. To directly test this hypothesis, we used a bar-coded transposon insertion library in tandem with cell sorting to assess genome-wide impact of gene deletions on membrane protein expression. The expression of five membrane proteins (CyoB, CydB, MdlB, YidC, and LepI) and one soluble protein (GST), each fused to GFP, was examined. We identified Escherichia coli mutants that demonstrated increased membrane protein expression relative to that in wild type. For two of the proteins (CyoB and CydB), we conducted functional assays to confirm that the increase in protein expression also led to phenotypic improvement in function. This study represents a systematic approach to broadly identify genetic loci that can be used to improve membrane protein expression, and our method can be used to improve expression of any protein that poses a cellular burden.
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Gene Editing
/
Membrane Proteins
Type of study:
Prognostic_studies
Language:
En
Journal:
Sci Rep
Year:
2017
Document type:
Article
Affiliation country:
Estados Unidos