Document Detail

High-yield membrane protein expression from E. coli using an engineered outer membrane protein F fusion.
MedLine Citation:
PMID:  23345122     Owner:  NLM     Status:  MEDLINE    
Obtaining high yields of membrane proteins necessary to perform detailed structural study is difficult due to poor solubility and variability in yields from heterologous expression systems. To address this issue, an Escherichia coli-based membrane protein overexpression system utilizing an engineered bacterial outer membrane protein F (pOmpF) fusion has been developed. Full-length human receptor activity-modifying protein 1 (RAMP1) was expressed using pOmpF, solubilized in FC15 and purified to homogeneity. Using circular dichroism and fluorescence spectroscopy, purified full-length RAMP1 is composed of approximately 90% α-helix, and retains its solubility and structure in FC15 over a wide range of temperatures (20-60°C). Thus, our approach provides a useful, complementary approach to achieve high-yield, full-length membrane protein overexpression for biophysical studies.
Pin-Chuan Su; William Si; Deidre L Baker; Bryan W Berger
Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, Non-P.H.S.     Date:  2013-02-21
Journal Detail:
Title:  Protein science : a publication of the Protein Society     Volume:  22     ISSN:  1469-896X     ISO Abbreviation:  Protein Sci.     Publication Date:  2013 Apr 
Date Detail:
Created Date:  2013-03-20     Completed Date:  2013-08-26     Revised Date:  2014-04-01    
Medline Journal Info:
Nlm Unique ID:  9211750     Medline TA:  Protein Sci     Country:  United States    
Other Details:
Languages:  eng     Pagination:  434-43     Citation Subset:  IM    
Copyright Information:
Copyright © 2013 The Protein Society.
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MeSH Terms
Circular Dichroism
Escherichia coli / genetics,  metabolism*
Escherichia coli Proteins / biosynthesis,  chemistry,  genetics,  metabolism
Inclusion Bodies / chemistry,  metabolism
Porins / biosynthesis*,  chemistry,  genetics,  metabolism
Protein Engineering / methods
Receptor Activity-Modifying Protein 1 / biosynthesis*,  chemistry,  genetics,  metabolism
Recombinant Fusion Proteins / biosynthesis*,  chemistry,  genetics,  metabolism
Reg. No./Substance:
0/Escherichia coli Proteins; 0/OmpF protein; 0/Porins; 0/RAMP1 protein, human; 0/Receptor Activity-Modifying Protein 1; 0/Recombinant Fusion Proteins

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