Crystal structure of xylose isomerase from thermophilic bacteriaThermus caldophilus

Glucose oxidase.a highly glycosylated flavoprotein, is one of the most widely usee! enzymes in medical diagnostics and food processing.The enormous economic significance and intensive application of glucose oxidase as biosensor enzyme, as integral part of food production and in fermentation control was contradicted so far by the lack of structural knowledge.The enzyme is a homodimer of !55 Iilla with a glycan moiety of the 'high mannose' type representing 13% of the molecular weight.Crystals suitable for X-ray diffraction were obtained only after enzymatic cleglycosylation removing 95% of the sacchmide residues.As confirmed by the high resolution structure the monomer of the deglycosylated protein comprises 587 amino acid residues, 5 N-acetyl glucosamine.3 mannose and l FAD molecule.The exceptional stability of glucose oxidase might be based on the lm-ge surface area bmied upon dime1ization and defined by (i) hydrophobic contacts, (ii) extensive salt bridges and, surprisingly, (iii) by a cluster of well ordered water molecules trapped between the monomers with no contact to the bulk.The 67 lilla monomer is comprised of two domains only.The FAD binding domain is separated from the substrate binding domain by a deep cavity filled with a well defined network of l 0 water molecules.At the given resolution this water cluster might suit as a stm•ting point for the modeling of the substrate glucose bound to the binary enzyme:FAD complex.The high resolution structure of glucose oxidase facilitates the use of protein engineering with the goal to design highly active enzyme derivatives with the capability to transfer directly electrons to semi-conductors and/or increased tolerance against hydrogen peroxide, sulfur dioxide and hydrogen sulfite.PS04.01.