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A conserved region within the Bordetella pertussis autotransporter BrkA is necessary for folding of its passenger domain
Author(s) -
Oliver David C.,
Huang George,
Nodel Elena,
Pleasance Steve,
Fernandez Rachel C.
Publication year - 2003
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1046/j.1365-2958.2003.03377.x
Subject(s) - biology , secretion , chaperone (clinical) , bacterial outer membrane , mutant , escherichia coli , signal peptide , effector , bordetella pertussis , protein folding , transport protein , cytoplasm , microbiology and biotechnology , biochemistry , peptide sequence , bacteria , gene , genetics , medicine , pathology
Summary Autotransporter secretion represents a unique mechanism that Gram‐negative bacteria employ to deliver proteins to their cell surface. BrkA is a Bordetella pertussis autotransporter protein that mediates serum resistance and contributes to adherence of the bacterium to host cells. BrkA is a 103 kDa protein that is cleaved to form a 73 kDa α‐domain and a 30 kDa β domain. The α domain, also referred to as the passenger domain, is responsible for the effector functions of the protein, whereas the β domain serves as a transporter. In an effort to characterize BrkA secretion, we have shown that BrkA has a 42 amino acid signal peptide for transit across the cytoplasmic membrane, and a translocation unit made up of a short linker region fused to the β‐domain to ferry the passenger domain to the bacterial surface through a channel formed by the β‐domain. In this report, we provide genetic, biochemical and structural evidence demonstrating that a region within the BrkA passenger (Glu 601 –Ala 692 ) is necessary for folding the passenger. This region is not required for surface display in the outer membrane protease OmpT‐deficient Escherichia coli strain UT5600. However, a BrkA mutant protein bearing a deletion in this region is susceptible to digestion when expressed in E. coli strains expressing OmpT suggesting that the region is required to maintain a stable structure. The instability of the deletion mutant can be rescued by surface expressing Glu 601 –Ala 692 in trans suggesting that this region is acting as an intramolecular chaperone to effect folding of the passenger concurrent with or following translocation across the outer membrane.