Figure 3.

(A) Thumbnail representation of the Bcs secretion macrocomplex components in E. coli-like (Type II) cellulose secretion systems (Krasteva et al. 2017; Zouhir et al. 2020; Abidi et al. 2021). (B) Cryo-EM structure of the assembled BcsRQABEF macrocomplex (emd-11836) showing hexameric BcsB in the crown (based on local refinement of pentameric BcsB^peri; average resolution 2.9 Å; accessions pdb-6yg8 and emd-10799) (Abidi et al. 2021). The BcsA subunit was modeled in Robetta and refined in the experimental BcsRQAB electron density following local refinement (emd-11836). The crystal structure of a BcsRQ complex was rigid-body fitted in the apical density. A predicted model for a dimer of BcsE's N-terminal domains (NTD, in cyan) was fitted in the bilobal membrane-proximal densities opposite BcsA based on fold prediction in Robetta, reported head-to-tail BcsE^NTD oligomerization and interactions with inner-membrane BcsF (Zouhir et al. 2020; Abidi et al. 2021). (C) A locally refined atomic model of the BcsRQAB assembly as found in the BcsRQABEF macrocomplex (emd-11836). An ∼140-residue-long N-terminal BcsA domain remains unresolved in the structure (Abidi et al. 2021). (D) Cryo-EM structure of purified full-length E. coli BcsB^His (emd-11356) with two octamers of BcsB's periplasmic domains refined in the experimental electron density (Abidi et al. 2021). Comparison of the E. coli and R. sphaeroides BcsB in the context of the BcsAB complex. (E) 1:1 BcsAB assemblies are shown with BcsA in white surface representation. Co-crystallized or modeled cellulose is shown as sticks. BcsB homologs are presented in cartoons with separate domains color-coded as follows: D1, blue; D2, cyan; D3, green; D4, yellow; C-terminal amphipathic and transmembrane helices, red and bordeaux, respectively. The R. sphaeroides D4 amphipathic helix insertion, substituted by an intersubunit 3-stranded β-sheet in E. coli BcsB^D4, is colored in orange. (F) Atomic model in surface representation of the E. coli BcsAB₆ assembly as found in the BcsRQABEF macrocomplex. Inset, β-sheet complementation between the D2′ and D4 flavodoxin-like modules from neighboring BcsB protomers, with the D2′ β-strand extension in dark blue and the D4 3-stranded β-sheet in orange. (G) Top view of the BcsB crown hexamer; domain color coding as in panel (E). Stacked D3 luminal loops are shown in transparent surface representation. Nascent cellulose (illustrated as stacked hexagons) is proposed to be extruded along the crown lumen with D3 luminal loops providing a ratchet-like structural support (Abidi et al. 2021).