Structural and biochemical investigation of PglF from Campylobacter jejuni reveals a new mechanism for a member of the short chain dehydrogenase/reductase superfamily
| Autor: | James B. Thoden, Ian C. Schoenhofen, N. Martin Young, Peter A. Tipton, David C. Watson, Alexander S. Riegert, Hazel M. Holden |
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| Jazyk: | angličtina |
| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
Reductase Crystallography X-Ray Biochemistry Campylobacter jejuni Article Catalysis Substrate Specificity 03 medical and health sciences chemistry.chemical_compound Bacterial Proteins Biosynthesis Cloning Molecular Genetics chemistry.chemical_classification Short-chain dehydrogenase Methionine 030102 biochemistry & molecular biology biology biology.organism_classification carbohydrates (lipids) Kinetics 030104 developmental biology Enzyme chemistry Membrane protein NAD+ kinase Oxidoreductases |
| Popis: | Within recent years it has become apparent that protein glycosylation is not limited to eukaryotes. Indeed, in Campylobacter jejuni, a Gram-negative bacterium, more than 60 of its proteins are known to be glycosylated. One of the sugars found in such glycosylated proteins is 2,4-diacetamido-2,4,6-trideoxy-α-d-glucopyranose, hereafter referred to as QuiNAc4NAc. The pathway for its biosynthesis, initiating with UDP-GlcNAc, requires three enzymes referred to as PglF, PglE, and PlgD. The focus of this investigation is on PglF, an NAD+-dependent sugar 4,6-dehydratase known to belong to the short chain dehydrogenase/reductase (SDR) superfamily. Specifically, PglF catalyzes the first step in the pathway, namely, the dehydration of UDP-GlcNAc to UDP-2-acetamido-2,6-dideoxy-α-d-xylo-hexos-4-ulose. Most members of the SDR superfamily contain a characteristic signature sequence of YXXXK where the conserved tyrosine functions as a catalytic acid or a base. Strikingly, in PglF, this residue is a methionine. Here we describe a detailed structural and functional investigation of PglF from C. jejuni. For this investigation five X-ray structures were determined to resolutions of 2.0 Å or better. In addition, kinetic analyses of the wild-type and site-directed variants were performed. On the basis of the data reported herein, a new catalytic mechanism for a SDR superfamily member is proposed that does not require the typically conserved tyrosine residue. |
| Databáze: | OpenAIRE |
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