CRYSTAL-STRUCTURES OF 3 COMPLEXES BETWEEN CHITO-OLIGOSACCHARIDES AND LYSOZYME FROM THE RAINBOW-TROUT - HOW DISTORTED IS THE NAG SUGAR IN SITE-D

Like all c-type lysozymes, those from rainbow trout act as 1,4-beta-ac etyl-muramidases to destroy bacteria by cleaving the polysaccharide ch ains of alternating N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM) units in the cell walls. Lysozymes also hydrolyse chitin, the a nalogous N-acetylglucosamine polymer. The rainbow trout enzymes have b een shown to be particularly effective in bacterial defence. We have d etermined the crystal structures of three complexes between rainbow tr out lysozyme (RBTL) and the chito-oligosaccharides (NAG)(2), (NAG)(3) and (NAG)(4) to resolutions of 1.8, 2.0 and 1.6 Angstrom, respectively . Crystals of these complexes were obtained by co-crystallization, and intensity data were collected on a FAST area detector system. Refinem ent and model building gave final R values of 16.6, 15.9 and 16.5% for the di-, tri- and tetrasaccharide complexes, respectively. The result s show that the chito-oligosaccharides bind to sites A, B and C as pre viously observed for complexes between the hen egg-white lysozyme (HEW L) and a variety of saccharides. The NAG ring in site D is not bound s o deeply and is only slightly distorted towards a half-chair conformat ion as observed for the equivalent NAM residue in HEWL. From our resul ts, there is reason to question the position and the degree of strain of the D saccharide and the mode of binding and importance of two sacc harides in sites E and F for correct orientation of sugar D and effect ive hydrolysis of a productive substrate-lysozyme complex. Simple mode l building study from our structures implies a 'left-sided' binding mo de of (NAG)(6) in the lower part of the active site of RBTL.