Alternative substrate-bound conformation of bacterial solute-binding protein involved in the import of mammalian host glycosaminoglycans

Sayoko Oiki,Reiko Kamochi,Wataru Hashimoto,Bunzo Mikami,Kousaku Murata

doi:10.1038/s41598-017-16801-8

Sayoko Oiki, Reiko Kamochi + Show 3 more

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https://doi.org/10.1038/s41598-017-16801-8

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Journal: Scientific Reports	Publication Date: Dec 1, 2017
Citations: 7	License type: open-access

Affiliation: Kyoto University, Setsunan University

Abstract

Glycosaminoglycans (GAGs), constituted by repeating uronate and amino sugar units, are major components of mammalian extracellular matrices. Some indigenous and pathogenic bacteria target GAGs for colonization to and/or infection of host mammalian cells. In Gram-negative pathogenic Streptobacillus moniliformis, the solute-binding protein (Smon0123)-dependent ATP-binding cassette (ABC) transporter incorporates unsaturated GAG disaccharides into the cytoplasm after depolymerization by polysaccharide lyase. Smon0123, composed of N and C domains, adopts either a substrate-free open or a substrate-bound closed form by approaching two domains at 47° in comparison with the open form. Here we show an alternative 39°-closed conformation of Smon0123 bound to unsaturated chondroitin disaccharide sulfated at the C-4 and C-6 positions of N-acetyl-d-galactosamine residue (CΔ4S6S). In CΔ4S6S-bound Smon0123, Arg204 and Lys210 around the two sulfate groups were located at different positions from those at other substrate-bound 47°-closed conformations. Therefore, the two sulfate groups in CΔ4S6S shifted substrate-binding residue arrangements, causing dynamic conformational change. Smon0123 showed less affinity with CΔ4S6S than with non-sulfated and monosulfated substrates. ATPase activity of the Smon0123-dependent ABC transporter in the presence of CΔ4S6S was lower than that in the presence of other unsaturated chondroitin disaccharides, suggesting that CΔ4S6S-bound Smon0123 was unpreferable for docking with the ABC transporter.

Highlights

Extracellular matrices are fibrillar network structures that lie under epithelial tissue cells and surround connective tissue cells[1]
Depending on the position and/or level of the sulfate groups, chondroitin sulfate is divided to several groups such as chondroitin sulfates A and B with a sulfate group at the C-4 position of GalNAc; chondroitin sulfate C with a sulfate group at the C-6 position of GalNAc; chondroitin sulfate D with two sulfate groups at the C-2 position of glucuronic acid (GlcUA) and the C-6 position of GalNAc; and chondroitin sulfate E sulfated at the C-4 and C-6 positions of GalNAc12
Bacterial cells were cultured on a plate that contained bovine serum albumin (BSA) and chondroitin sulfate A

Summary

Introduction

Extracellular matrices are fibrillar network structures that lie under epithelial tissue cells and surround connective tissue cells[1]. Enzymes, i.e. isomerase, NADH-dependent reductase, kinase, and aldolase, subsequently metabolize the unsaturated uronate to pyruvate and glyceraldehyde-3-phosphate[7] The genes encoding these enzymes that are essential for depolymerization, degradation, and metabolism of GAGs assemble a cluster in the bacterial genome. This study assesses the characteristic conformational change of CΔ4S6S-bound Smon0123, the structural determinants of Smon0123 for substrate-dependent conformations, and the modeling of Smon0123/Smon0121-Smon0122/ Smon0120-Smon0120 on the basis of the alginate ABC transporter system (AlgQ2/AlgM1-AlgM2/AlgS-AlgS). These findings provide structural and functional insights for further understanding substrate recognition and import and for developing inhibitors for pathogenic bacteria

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