Bornemann S, Ramjee M K, Balasubramanian S, Abell C, Coggins J R, Lowe D J, Thorneley R N
Nitrogen Fixation Laboratory, University of Sussex, Brighton, United Kingdom.
J Biol Chem. 1995 Sep 29;270(39):22811-5. doi: 10.1074/jbc.270.39.22811.
Chorismate synthase catalyzes the conversion of 5-enolpyruvylshikimate-3-phosphate to chorismate. It is the seventh enzyme of the shikimate pathway, which is responsible for the biosynthesis of aromatic metabolites from glucose. The chorismate synthase reaction involves a 1,4-elimination with unusual anti-stereochemistry and requires a reduced flavin cofactor. The substrate analogue (6S)-6-fluoro-5-enolpyruvylshikimate-3-phosphate is a competitive inhibitor of Neurospora crassa chorismate synthase (Balasubramanian, S., Davies, G. M., Coggins, J. R., and Abell, C. (1991) J. Am. Chem. Soc. 113, 8945-8946). We have shown that this analogue is converted to 6-fluorochorismate by Escherichia coli chorismate synthase at a rate 2 orders of magnitude slower than the normal substrate. The decreased rate of reaction is consistent with the destabilization of an allylic cationic intermediate. The formation of chorismate and 6-fluorochorismate involves a common protein-bound flavin intermediate although the fluoro substituent does influence the spectral characteristics of this intermediate. The fluoro substituent also decreased the rate of decay of the flavin intermediate by 280 times. These results are consistent with the antimicrobial activity of (6S)-6-fluoroshikimate not being mediated by the inhibition of chorismate synthase but by the inhibition of 4-aminobenzoic acid synthesis as previously proposed (Davies, G. M., Barrett-Bee, K. J., Jude, D. A., Lehan, M., Nichols, W. W., Pinder, P. E., Thain, J. L., Watkins, W. J., and Wilson, R. G. (1994) Antimicrobial Agents and Chemotherapy 38, 403-406).
分支酸合酶催化5-烯醇丙酮酸莽草酸-3-磷酸转化为分支酸。它是莽草酸途径的第七种酶，该途径负责从葡萄糖生物合成芳香族代谢物。分支酸合酶反应涉及具有异常反式立体化学的1,4-消除反应，并且需要还原型黄素辅因子。底物类似物(6S)-6-氟-5-烯醇丙酮酸莽草酸-3-磷酸是粗糙脉孢菌分支酸合酶的竞争性抑制剂（巴拉苏布拉马尼亚姆，S.，戴维斯，G.M.，科金斯，J.R.，和阿贝尔，C.（1991年）《美国化学会志》113，8945 - 8946）。我们已经表明，大肠杆菌分支酸合酶将这种类似物转化为6-氟分支酸的速率比正常底物慢2个数量级。反应速率的降低与烯丙基阳离子中间体的不稳定相一致。分支酸和6-氟分支酸的形成涉及一种共同的蛋白质结合黄素中间体，尽管氟取代基确实会影响该中间体的光谱特征。氟取代基还使黄素中间体的衰变速率降低了280倍。这些结果与(6S)-6-氟莽草酸的抗菌活性不是由分支酸合酶的抑制介导的，而是如先前所提出的由4-氨基苯甲酸合成的抑制介导的观点一致（戴维斯，G.M.，巴雷特-比，K.J.，裘德，D.A.，莱汉，M.，尼科尔斯，W.W.，平德，P.E.，塞恩，J.L.，沃特金斯，W.J.，和威尔逊，R.G.（1994年）《抗菌剂与化疗》38，403 - 406）。
