Open Access
Crystallization and atomic resolution X‐ray diffraction of the catalytic domain of the large sialidase, nanI, from Clostridium perfringens
Author(s) -
Newstead Simon,
Chien ChinHsiang,
Taylor Margaret,
Taylor Garry
Publication year - 2004
Publication title -
acta crystallographica section d
Language(s) - English
Resource type - Journals
ISSN - 1399-0047
DOI - 10.1107/s090744490402181x
Subject(s) - sialidase , clostridium perfringens , glycan , escherichia coli , microbiology and biotechnology , clostridium , glycoprotein , biochemistry , enzyme , biology , sialic acid , glycosylation , glycolipid , bacteria , molecular replacement , chemistry , neuraminidase , gene , genetics
Sialidases catalyse the removal of terminal sialic acids from a range of glycoproteins, glycolipids and oligosaccharides. They have been found in bacteria, viruses and parasites, where they play important roles in pathogenesis and/or microbial nutrition, and in mammalian cells, where they modulate cell‐surface glycosylation associated with a range of cellular activities. Clostridium perfringens , a causative agent of gas gangrene and peritonitis in humans, possesses three sialidases: nanH, nanI and nanJ, with molecular weights of 42, 77 and 129 kDa, respectively. The two larger enzymes are secreted by the bacterium and are involved in the pathogenesis and nutrition of Clostridium . As part of a study to examine the structures of all three enzymes, crystallization of the 77 kDa nanI isoenzyme was attempted. The expressed full‐length protein was found to degrade easily; a stable 50 kDa catalytic domain was therefore subcloned. This domain was overexpressed in Escherichia coli and produced crystals belonging to space group P 2 1 2 1 2 1 , with unit‐cell parameters a = 96.98, b = 69.41, c = 72.69 Å and one monomer per asymmetric unit. The crystals diffract to at least 0.92 Å. A molecular‐replacement solution was obtained using the catalytic domain of the sialidase from the leech Macrobdella decora .