An electrophilic cyanation of aryl Grignard or lithium reagents, generated in situ from the corresponding aryl bromides or iodides, by a transnitrilation with dimethylmalononitrile (DMMN) was developed. DMMN is a commercially available, bench-stable solid. The transnitrilation with DMMN avoids the use of toxic reagents and transition metals and occurs under mild reaction conditions, even for extremely sterically hindered substrates. The transnitrilation of aryllithium species generated by directed ortho-lithiation enabled a net C-H cyanation. The intermediacy of a Thorpe-type imine adduct in the reaction was supported by isolation of the corresponding ketone from the quenched reaction. Computational studies supported the energetic favorability of retro-Thorpe fragmentation of the imine adduct.
Substrate analogues were synthesized and examined as inhibitors of diaminopimelic acid (DAP) dehydrogenase from Bacillus sphaericus and of DAP epimerase from Escherichia coli. These enzymes produce meso-DAP (3) (a precursor for L-lysine and for peptidoglycan) from L-tetrahydrodipicolinic acid (1) and ll-DAP (2), respectively. The epimerase was purified by an improved procedure and confirmed to require both carboxyl and both amino groups for substrate recognition using deuterium-exchange experiments with DAP isomers, L-lysine, D-lysine, L-a-aminopimelate, and racemic -aminopimelate. An imidazole analogue of DAP, (2S)-2-amino-3-(4-carboxyimidazol-1 -yl)propanoic acid (4), was synthesized by condensation of benzyl imidazole-4-carboxylate (8) with JV-benzyloxycarbonyl(Cbz)-Lserine jS-lactone (9) (product structure confirmed by X-ray analysis) followed by hydrogenolytic deprotection. Two other analogues, (2S,J'/?)-2-amino-3-(3-carboxy-2-isoxazolin-5-yl)propanoic acid (5) and its 5'S diastereomer 6, were prepared by condensation of methyl jV-Cbz-L-allylglycinate (13) with methyl chlorooximidoacetate ( 14) followed by separation of isomers and deprotection with N aOH and MeaSiCl/Nal. Similar condensation of ethyl chlorooximidoacetate with ethylene and of 14 with ethyl acrylate generated isoxazolines, which were saponified to 2-isoxazoline-3-carboxylic acid (25) and 2-isoxazoline-3,5-dicarboxylic acid (26), respectively. None of the compounds show significant inhibition of DAP epimerase or DAP dehydrogenase with the exception of 6, which is a potent and specific inhibitor of DAP dehydrogenase. At pH 7.5 or 7.8, compound 6 shows competitive inhibition (X¡ = 4.2 µ ) with tetrahydrodipicolinic acid (1) for the forward reaction and noncompetive inhibition (X¡ = 23 µ ) with meso-(3) for the reverse process.Preliminary tests for antimicrobial activity demonstrate that 6 inhibits the growth of B. sphaericus, which relies exclusively on DAP dehydrogenase to produce 3.
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