The site-directed mutagenesis of a number of proposed active site residues of 5-enolpyruvyl shikimate-3-phosphate (EPSP) synthase is reported. Several of these mutations resulted in complete loss of enzyme activity indicating that these residues are probably involved with catalysis, notably K22R, K411R, D384A, R27A, R100A, and D242A. Of those, K22R, R27A, and D384A did not bind either the substrate shikimate-3-phosphate (S3P) or glyphosate (GLP). The K411R and D242A mutants bind S3P only in the presence of GLP. The kinetic characterization of mutants R100K, K340R, and E418A, which retain activity, is reported. Of those, R100K and K340R do not accumulate enzyme intermediate of enzyme-bound product under equilibrium conditions. These residues, while not essential for catalysis, are most likely important for substrate binding. All of the mutants are shown to be correctly folded by NMR spectroscopy.

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Synthesis and Structure of Functionalized Derivatives of the Cleft-Shaped Molecule Dithiosalicylide.-A recently developed simple preparation of dithiosalicylide from 3H-1,2-benzodithiol-3-one is applied to the preparation of analogues presenting larger cavities than the parent compound and to several analogues which may be amenable to further functionalization. The crystal structures of two compounds show that they form discrete, selfincluded dimers and that interaction between the dimers give rise to interesting supermolecular structures. Because the dithiosalicylide-forming reaction involves dimerization of triphenylphosphine-activated 3H-1,2-benzodithiol-3-ones, it is possible to prepare asymmetric functionalized clefts by this method, starting with 1:1 mixtures of two different derivatives of (III). -(MITRA, K.; POHL, M. E.; MACGILLIVRAY, L. R.; BARNES, C. L.; GATES, K. S.; J. Org. Chem. 62 (1997) 26, 9361-9364; Dep. Chem., Univ. Mo., Columbia, MO 65211, USA; EN)