Structural and biochemical features distinguish the foot-and-mouth disease virus leader proteinase from other papain-like enzymes

The structures of the two leader protease (L-pro) variants of foot-and-mouth disease virus known to date were solved using crystals in which molecules were organized as molecular fibers. Such crystals diffract to a resolution of only approximately 3 Angstrom. This singular, pseudo-polymeric organization is present in a new L-pro crystal form showing a cubic packing. As molecular fiber formation appeared unrelated to crystallization conditions, we mutated the reactive cysteine 133 residue, which makes a disulfide bridge between adjacent monomers in the fibers, to serine. None of the intermolecular contacts found in the molecular fibers was present in crystals of this variant. Analysis of this L-pro structure, refined at 1.9 Angstrom resolution, enables a detailed definition of the active center of the enzyme, including the solvent organization. Assay of L-pro activity on a fluorescent hexapeptide substrate showed that L-pro, in contrast to papain, was highly sensitive to increases in the cation concentration and was active only across a narrow pH range. Examination of the L-pro structure revealed that three aspartate residues near the active site, not present in papain-like enzymes, are probably responsible for these properties. (C) 2000 Academic Press.