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Cooperative binding of LysM domains determines the carbohydrate affinity of a bacterial endopeptidase protein
Author(s) -
Wong Jaslyn E. M. M.,
Alsarraf Husam M. A. B.,
Kaspersen Jørn Døvling,
Pedersen Jan Skov,
Stougaard Jens,
Thirup Søren,
Blaise Mickaël
Publication year - 2014
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.12698
Subject(s) - autolysin , peptidoglycan , biochemistry , lysin , cell wall , cooperativity , chitin , bacillus subtilis , carbohydrate binding module , endopeptidase , chemistry , bacterial cell structure , microscale thermophoresis , biology , enzyme , glycoside hydrolase , bacteria , escherichia coli , gene , genetics , bacteriophage , chitosan
Cellulose, chitin and peptidoglycan are major long‐chain carbohydrates in living organisms, and constitute a substantial fraction of the biomass. Characterization of the biochemical basis of dynamic changes and degradation of these β,1–4‐linked carbohydrates is therefore important for both functional studies of biological polymers and biotechnology. Here, we investigated the functional role of multiplicity of the carbohydrate‐binding lysin motif ( L ys M ) domain that is found in proteins involved in bacterial peptidoglycan synthesis and remodelling. The B acillus subtilis peptidoglycan‐hydrolysing N lp C / P 60 d , l ‐endopeptidase, cell wall‐lytic enzyme associated with cell separation, possesses four L ys M domains. The contribution of each L ys M domain was determined by direct carbohydrate‐binding studies in aqueous solution with microscale thermophoresis. We found that bacterial L ys M domains have affinity for N‐acetylglucosamine (GlcNac) polymers in the lower‐micromolar range. Moreover, we demonstrated that a single L ys M domain is able to bind carbohydrate ligands, and that L ys M domains act additively to increase the binding affinity. Our study reveals that affinity for G lc NA c polymers correlates with the chain length of the carbohydrate, and suggests that binding of long carbohydrates is mediated by L ys M domain cooperativity. We also show that bacterial L ys M domains, in contrast to plant L ys M domains, do not discriminate between G lc NA c polymers, and recognize both peptidoglycan fragments and chitin polymers with similar affinity. Finally, an Ala replacement study suggested that the carbohydrate‐binding site in L ys M ‐containing proteins is conserved across phyla.