Effective assembly of fimbriae in Escherichia coli depends on the translocation assembly module nanomachine

Abstract

Character count:Current: 171 words in first paragraph 1497 words in rest of text (excluding methods, references, legends).Permitted: 200 words (300 words max) for first paragraph 1500 words max for rest (excluding methods, references, legends). The crystal structure of the fimbrial usher protein FimD revealed an intricate, five-domain architecture: a β-barrel domain, incorporated into which is a β-sandwich plug domain, an Nterminal periplasmic domain (NTD) and two C-terminal periplasmic domains (CTD1 and CTD2) 6,7 ( Supplementary Fig. S1). The mechanism for folding and assembling such a large, multi-domains, protein into the outer membrane is unclear, and currently no assay system is available to study the process in vivo. A candidate to catalyze this assembly reaction was the β-barrel assembly machinery (BAM complex): composed of the essential proteins BamA and BamD, assisted by the proteins BamB, BamC and BamE; Fig. 1a. In addition, bacteria haveTamA and TamB (Fig. 1a) that constitute a translocation and assembly module (TAM) [10][11][12] . The

Structural basis for substrate selection by the translocation and assembly module of the β‐barrel assembly machinery

“…In terms of stable interactions with the transmembrane β‐strand peptides, the barrel domains of both BamA and TamA are capable of mediating interactions, and they do so with high selectivity for their substrate. This observation begins to explain the rapid, and directional, folding of proteins like FimD mediated by the TAM (Stubenrauch et al ., ). TamA demonstrated a selectivity for elements of FimD over and above any binding observed for BamA.…”

An investigation into the Omp85 protein BamK in hypervirulent Klebsiella pneumoniae, and its role in outer membrane biogenesis

“…The assembly of trimeric porins can be measured with a pulse‐chase assay based on semi‐native PAGE to separate the assembled trimers from the monomeric forms of unassembled protein (Heinz et al, , Stubenrauch et al, ). To monitor the rate of assembly of the trimeric porin, translation of [ 35 S]‐labeled PhoE precursor was induced in each strain and the assembly of the porin trimer was determined by semi‐native PAGE and densitometry.…”

“…To monitor the rate of assembly of the trimeric porin, translation of [ 35 S]‐labeled PhoE precursor was induced in each strain and the assembly of the porin trimer was determined by semi‐native PAGE and densitometry. In the semi‐native gel electrophoresis the monomeric PhoE species is better focused than the oligomeric species, so the exponential decay of the monomer (as it assembles into an oligomer) was calculated (Stubenrauch et al, ) for the time‐course of PhoE assembly in E. coli Δ bamA :: KpbamA (Fig. C) and Δ bamA :: KpbamK (Fig.…”

“…LPS display on the cell surface depends on the activity of the LptDE complex, which is assembled by the BAM complex. The type I and type III fimbrial adhesins are extended through usher proteins (Klemm & Schembri, , Wilksch et al, , Khater et al, ), which are themselves dependent on the BAM complex and the TAM for efficient assembly (Stubenrauch et al, , Stubenrauch et al, , Heinz et al, ). While the Wza secretion pore for capsular polysaccharide is not assembled by the BAM complex (Dunstan et al, ), the capsule has a key attachment factor Wzi that is a β‐barrel protein (Bushell et al, ).…”

“…The Wza capsule secretion pore appears to self‐assemble in the outer membrane (Dunstan et al, ), whereas most other outer membrane proteins are assembled by a highly sophisticated β‐barrel assembly machinery (BAM) composed of the core BAM complex (BAM ABCDE ) and the translocation and assembly module (the TAM) (Webb et al, , Plummer & Fleming, , Albenne & Ieva, , Noinaj et al, ). Each of these protein complexes is constituted around a membrane protein of the Omp85 protein family (BamA and TamA), and both are involved in the assembly of complex structures such as the adhesive fimbriae (Stubenrauch et al, , Stubenrauch et al, ) required by K. pneumoniae for biofilm formation (Klemm & Schembri, , Wilksch et al, , Khater et al, , Tan et al, ).…”