In a similar fashion, bis(triphenylphosphine)palladium(II) chloride (3.64 g, 5.17 mmol, 0.050 equiv) was added (at 0 oC) and the resulting mixture was deoxygenated once again at-78 oC. The deoxygenated reaction suspension was then warmed to 23 oC and was stirred at that temperature for 3 h. The brown product solution was washed with a 1:1 mixture of saturated aqueous potassium carbonate solution and saturated aqueous ammonium chloride solution (3 x 150 mL). The organic layer was dried over sodium sulfate and was concentrated. The residue was purified by distillation under reduced pressure (bp 45-50 oC, 0.5 mm Hg) to afford (Z)-1-chloro-4-(tertbutyldimethylsilyl)-1-buten-3-yne (32) as a colorless oil (12.5 g, 60%

Three enzymes of the shikimic acid pathway, isochorismate synthase (IS), anthranilate synthase (AS), and p-aminobenzoate synthase (PABS), exhibit significant sequence homology and may be related mechanistically.Compounds 1, 2, and 3 were designed to mimic, in their all-axial conformations, the putative transition state for these enzymes. The inhibitors were prepared in racemic form starting from Diels-Alder addition of a propiolate ester to a protected 1-oxy-or 1-amino-1,3-butadiene in 14%, 4%, and 9% overall yields, respectively. All three compounds are competitive inhibitors of the three enzymes, binding IS and AS strongly and PABS weakly. For both IS and AS, the affinity of the 6-amino-4-hydroxy isomer 2 is ca. 10-fold that of the 4-amino-6-hydroxy isomer 3, a difference that is largely due to their conformational equilibria; 2 is 25 ± 2% axial and 3 is 6 ± 3% axial, as determined by the temperature dependence of their NMR spectra. The similarity between IS and AS was extended by the finding that IS, like AS, catalyzes formation of 2-amino-2-deoxyisochorismate (ADIC) in the presence of ammonia. These observations are consistent with direct 1,5-substitution mechanisms for both IS and AS; the weak inhibition of PABS by these inhibitors suggests that it operates by a significantly different mechanism.

VanA has greatly reduced affinity for all the ligands studied. The relative affinities of the inhibitors in the reversible binding step are not, however, consistent with the substrate specificities of the enzymes. We propose a mechanism in which proton transfer from the attacking nucleophile to the departing phosphate occurs directly, without intervention of the enzyme.

Enantiomerically pure dynemicin A is now available by laboratory synthesis. The natural, (+)-enantiomer of dynemicin A is shown to possess the 2S, 3S, 4S, 7R, 8R configuration. A wide variety of heretofore unavailable, active analogs of dynemicin A have been prepared and are found to produce subtle variations in sequence specificity of DNA cleavage compared to the natural product and, of potentially greater significance, display variations in the efficiency of DNA cleavage as a function of the activating agent.

No abstract