Abstract
Bacteria produce a remarkably diverse range of glycoside hydrolases to metabolize glycans from the environment as a primary source of nutrients, and to promote the colonization and infection of a host. Here we focus on EndoE, a multi-modular glycoside hydrolase secreted by Enterococcus faecalis, one of the leading causes of healthcare-associated infections. We provide X-ray crystal structures of EndoE, which show an architecture composed of four domains, including GH18 and GH20 glycoside hydrolases connected by two consecutive three α-helical bundles. We determine that the GH20 domain is an exo-β-1,2-N-acetylglucosaminidase, whereas the GH18 domain is an endo-β-1,4-N-acetylglucosaminidase that exclusively processes the central core of complex-type or high-mannose-type N-glycans. Both glycoside hydrolase domains act in a concerted manner to process diverse N-glycans on glycoproteins, including therapeutic IgG antibodies. EndoE combines two enzyme domains with distinct functions and glycan specificities to play a dual role in glycan metabolism and immune evasion.
Highlights
Bacteria produce a remarkably diverse range of glycoside hydrolases to metabolize glycans from the environment as a primary source of nutrients, and to promote the colonization and infection of a host
EndoE-GH18L crystallized in the P 6 5 space group with one molecule in the asymmetric unit and diffracted to a maximum resolution of 1.7 Å
The study of the molecular mechanism of substrate recognition of the individual domains of multi-modular glycoside hydrolases such as EndoE and how these domains can work synergistically is of critical importance to understand their mode of action in the context of a particular biological function and to the rational design of novel enzymes that can optimize the glycoengineering of glycoproteins
Summary
Bacteria produce a remarkably diverse range of glycoside hydrolases to metabolize glycans from the environment as a primary source of nutrients, and to promote the colonization and infection of a host. We determine that the GH20 domain is an exo-β-1,2-N-acetylglucosaminidase, whereas the GH18 domain is an endo-β-1,4-N-acetylglucosaminidase that exclusively processes the central core of complex-type or high-mannose-type N-glycans. Both glycoside hydrolase domains act in a concerted manner to process diverse N-glycans on glycoproteins, including therapeutic IgG antibodies. Glycans play a central role in bacterial physiology and pathogenicity They are used primarily as structural constituents of bacterial cell envelopes as well as metabolic intermediates and molecules that allow energy storage. Glycan modification of proteins and lipids generates substantial structural diversity in bacteria These structures play critical roles in molecular recognition events including the evasion of the immune response and hostpathogen interactions. Glycans are broken down and released from carbohydrate and non-carbohydrate moieties into their constituent monosaccharides by enzymes known as glycoside hydrolases (GHs)
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