Abstract

Bacterial outer membrane proteins (OMPs) contain a unique “β barrel” segment that is inserted into the membrane by the barrel assembly machinery (Bam) complex by an unknown mechanism. OMP assembly has been reconstituted in vitro, but assembly reactions have involved the use of urea-denatured protein purified from inclusion bodies. Here we show that the E. coli Bam complex catalyzes the efficient assembly of OMPs synthesized de novo in a coupled in vitro transcription/translation system. Interestingly, the in vitro translated forms of the OMPs we analyzed were assembled more rapidly and were effectively engaged by fewer periplasmic chaperones than their urea-denatured counterparts. Taken together, our results strongly suggest that the mode of production influences the conformational states sampled by OMPs and thereby affects their recognition by both chaperones and the Bam complex. Besides providing insights into OMP biogenesis, our work describes a novel, streamlined method to reconstitute OMP assembly in vitro.

Details

Title
Bam complex-mediated assembly of bacterial outer membrane proteins synthesized in an in vitro translation system
Author
Hussain Sunyia 1 ; Peterson, Janine H 2 ; Bernstein, Harris D 2 

 Genetics and Biochemistry Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, USA (GRID:grid.419635.c) (ISNI:0000 0001 2203 7304); TetraGenetics, Inc., 91 Mystic St., Arlington, USA (GRID:grid.479566.f) 
 Genetics and Biochemistry Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, USA (GRID:grid.419635.c) (ISNI:0000 0001 2203 7304) 
Publication year
2020
Publication date
2020
Publisher
Nature Publishing Group
e-ISSN
20452322
Source type
Scholarly Journal
Language of publication
English
ProQuest document ID
2376714861
Copyright
This work is published under http://creativecommons.org/licenses/by/4.0/ (the “License”). Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.