7RAW

Domain 1 of Starch adherence system protein 20 (Sas20) from Ruminococcus bromii

Summary for 7RAW

• Entry DOI: 10.2210/pdb7raw/pdb
• Descriptor: Dockerin domain-containing protein, DI(HYDROXYETHYL)ETHER, TRIETHYLENE GLYCOL, ... (4 entities in total)
• Functional Keywords: starch-binding protein domain in the ruminococcus bromii amylosome protein sas20, sugar binding protein
• Biological source: Ruminococcus bromii L2-63
• Total number of polymer chains: 1
• Total formula weight: 27780.60

Authors

Cerqueira, F.M.,Koropatkin, N.M. (deposition date: 2021-07-04, release date: 2022-04-13, Last modification date: 2024-05-22)

Primary citation

Cerqueira, F.M.,Photenhauer, A.L.,Doden, H.L.,Brown, A.N.,Abdel-Hamid, A.M.,Morais, S.,Bayer, E.A.,Wawrzak, Z.,Cann, I.,Ridlon, J.M.,Hopkins, J.B.,Koropatkin, N.M.
Sas20 is a highly flexible starch-binding protein in the Ruminococcus bromii cell-surface amylosome.
J.Biol.Chem., 298:101896-101896, 2022

PubMed Abstract: Ruminococcus bromii is a keystone species in the human gut that has the rare ability to degrade dietary resistant starch (RS). This bacterium secretes a suite of starch-active proteins that work together within larger complexes called amylosomes that allow R. bromii to bind and degrade RS. Starch adherence system protein 20 (Sas20) is one of the more abundant proteins assembled within amylosomes, but little could be predicted about its molecular features based on amino acid sequence. Here, we performed a structure-function analysis of Sas20 and determined that it features two discrete starch-binding domains separated by a flexible linker. We show that Sas20 domain 1 contains an N-terminal β-sandwich followed by a cluster of α-helices, and the nonreducing end of maltooligosaccharides can be captured between these structural features. Furthermore, the crystal structure of a close homolog of Sas20 domain 2 revealed a unique bilobed starch-binding groove that targets the helical α1,4-linked glycan chains found in amorphous regions of amylopectin and crystalline regions of amylose. Affinity PAGE and isothermal titration calorimetry demonstrated that both domains bind maltoheptaose and soluble starch with relatively high affinity (K ≤ 20 μM) but exhibit limited or no binding to cyclodextrins. Finally, small-angle X-ray scattering analysis of the individual and combined domains support that these structures are highly flexible, which may allow the protein to adopt conformations that enhance its starch-targeting efficiency. Taken together, we conclude that Sas20 binds distinct features within the starch granule, facilitating the ability of R. bromii to hydrolyze dietary RS.

PubMed: 35378131
DOI: 10.1016/j.jbc.2022.101896

Experimental method

X-RAY DIFFRACTION (2.1 Å)