Crystal Structure of a Bacterial Unsaturated Glucuronyl Hydrolase with Specificity for Heparin

Nakamichi, Yusuke; Mikami, Bunzo; Murata, Kousaku; Hashimoto, Wataru

doi:10.1074/jbc.m113.522573

Cited by 8 publications

(6 citation statements)

References 42 publications

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“…In addition, mutation of Pro44 to some hydrophilic amino acids increasing the activity of SulE may also be due to the alteration of the lid loop flexibility. Our finding is consistent with some previous reports of different types of enzymes; that is, the modulation of loop flexibilities could alter enzyme properties 37 – 39 .…”

Section: Discussionsupporting

confidence: 93%

Crystal structures of herbicide-detoxifying esterase reveal a lid loop affecting substrate binding and activity

et al. 2023

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SulE, an esterase, which detoxifies a variety of sulfonylurea herbicides through de-esterification, provides an attractive approach to remove environmental sulfonylurea herbicides and develop herbicide-tolerant crops. Here, we determined the crystal structures of SulE and an activity improved mutant P44R. Structural analysis revealed that SulE is a dimer with spacious binding pocket accommodating the large sulfonylureas substrate. Particularly, SulE contains a protruding β hairpin with a lid loop covering the active site of the other subunit of the dimer. The lid loop participates in substrate recognition and binding. P44R mutation altered the lid loop flexibility, resulting in the sulfonylurea heterocyclic ring repositioning to a relative stable conformation thus leading to dramatically increased activity. Our work provides important insights into the molecular mechanism of SulE, and establish a solid foundation for further improving the enzyme activity to various sulfonylurea herbicides through rational design.

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Section: Discussionsupporting

confidence: 93%

Crystal structures of herbicide-detoxifying esterase reveal a lid loop affecting substrate binding and activity

et al. 2023

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“…Unsaturated GAG disaccharides, ΔHA, CΔ0S, CΔ4S, CΔ6S, HΔ0S, and HΔ6S, were used as substrates. One unit of the enzyme activity was defined as the amount of enzyme required to degrade 1 µmol of substrate per minute as described previously 12 , 48 . The enzyme was also assayed using various concentrations (0 to 1 mM) of CΔ0S, and kinetic parameters ( K m and k cat ) were determined according the Michaelis-Menten equation (Synergy Software).…”

Section: Methodsmentioning

confidence: 99%

A bacterial ABC transporter enables import of mammalian host glycosaminoglycans

Oiki

Mikami

Maruyama

et al. 2017

Sci Rep

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Glycosaminoglycans (GAGs), such as hyaluronan, chondroitin sulfate, and heparin, constitute mammalian extracellular matrices. The uronate and amino sugar residues in hyaluronan and chondroitin sulfate are linked by 1,3-glycoside bond, while heparin contains 1,4-glycoside bond. Some bacteria target GAGs as means of establishing colonization and/or infection, and bacterial degradation mechanisms of GAGs have been well characterized. However, little is known about the bacterial import of GAGs. Here, we show a GAG import system, comprised of a solute-binding protein (Smon0123)-dependent ATP-binding cassette (ABC) transporter, in the pathogenic Streptobacillus moniliformis. A genetic cluster responsible for depolymerization, degradation, and metabolism of GAGs as well as the ABC transporter system was found in the S. moniliformis genome. This bacterium degraded hyaluronan and chondroitin sulfate with an expression of the genetic cluster, while heparin repressed the bacterial growth. The purified recombinant Smon0123 exhibited an affinity with disaccharides generated from hyaluronan and chondroitin sulfate. X-ray crystallography indicated binding mode of Smon0123 to GAG disaccharides. The purified recombinant ABC transporter as a tetramer (Smon0121-Smon0122/Smon0120-Smon0120) reconstructed in liposomes enhanced its ATPase activity in the presence of Smon0123 and GAG disaccharides. This is the first report that has molecularly depicted a bacterial import system of both sulfated and non-sulfated GAGs.

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“…Hep I lyases require sulfation, Hep II PLs exhibit promiscuity with respect to sulfation patterns, and Hep III enzymes cleave low-sulfation regions of these GAGs (13,14). The lyases generate products capped by Δ4,5-unsaturated UA, which is released from the GAG by GHs from family GH88 (13). In addition, some GAG-specific sulfatases have been characterized (15).…”

Section: Significancementioning

confidence: 99%

How members of the human gut microbiota overcome the sulfation problem posed by glycosaminoglycans

Cartmell

Lowe

Baslé

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

120

143

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The human microbiota, which plays an important role in health and disease, uses complex carbohydrates as a major source of nutrients. Utilization hierarchy indicates that the host glycosaminoglycans heparin (Hep) and heparan sulfate (HS) are high-priority carbohydrates for , a prominent member of the human microbiota. The sulfation patterns of these glycosaminoglycans are highly variable, which presents a significant enzymatic challenge to the polysaccharide lyases and sulfatases that mediate degradation. It is possible that the bacterium recruits lyases with highly plastic specificities and expresses a repertoire of enzymes that target substructures of the glycosaminoglycans with variable sulfation or that the glycans are desulfated before cleavage by the lyases. To distinguish between these mechanisms, the components of the Hep/HS degrading apparatus were analyzed. The data showed that the bacterium expressed a single-surface endo-acting lyase that cleaved HS, reflecting its higher molecular weight compared with Hep. Both Hep and HS oligosaccharides imported into the periplasm were degraded by a repertoire of lyases, with each enzyme displaying specificity for substructures within these glycosaminoglycans that display a different degree of sulfation. Furthermore, the crystal structures of a key surface glycan binding protein, which is able to bind both Hep and HS, and periplasmic sulfatases reveal the major specificity determinants for these proteins. The locus described here is highly conserved within the human gut , indicating that the model developed is of generic relevance to this important microbial community.

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Crystal Structure of a Bacterial Unsaturated Glucuronyl Hydrolase with Specificity for Heparin

Cited by 8 publications

References 42 publications

Crystal structures of herbicide-detoxifying esterase reveal a lid loop affecting substrate binding and activity

Crystal structures of herbicide-detoxifying esterase reveal a lid loop affecting substrate binding and activity

A bacterial ABC transporter enables import of mammalian host glycosaminoglycans

How members of the human gut microbiota overcome the sulfation problem posed by glycosaminoglycans

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