Phillips R S, Von Tersch R L, Secundo F
Department of Chemistry, University of Georgia, Athens 30602-2556, USA.
Eur J Biochem. 1997 Mar 1;244(2):658-63. doi: 10.1111/j.1432-1033.1997.00658.x.
The interactions of ring fluorinated analogs of tyrosine with tyrosine phenol-lyase and tryptophan indole-lyase (tryptophanase) were studied by rapid-scanning stopped-flow spectrophotometry. The reaction of L-tyrosine with tyrosine phenol-lyase resulted in rapid formation of a small absorbance peak at 500 nm, attributed to a quinonoid intermediate. The reaction of 3-fluoro-L-tyrosine with tyrosine phenol-lyase resulted in a peak at 500 nm with much higher absorbance, as did the reaction of 3,5-difluoro-L-tyrosine, due to increased accumulation of quinonoid intermediates. In constrast, complexes with 2-fluoro-L-tyrosine, 2,3-difluoro-L-tyrosine, 2,5-difluoro-L-tyrosine, and 2,6-difluoro-L-tyrosine exhibited much lower absorbance intensity at 500 nm. The rate constant for quinonoid intermediate formation from 3-fluoro-L-tyrosine was comparable to that for L-tyrosine. However, 3,5-difluoro-L-tyrosine reacted to form a quinonoid intermediate at about half the rate of L-tyrosine, while 2,3-difluoro-L-tyrosine reacted at twice the rate of L-tyrosine. In addition, the 2-substituted difluorotyrosines exhibited an intermediate, which was formed rapidly, absorbing strongly at about 340 nm, which is likely due to a gem-diamine intermediate. Tyrosine is not a substrate for tryptophan indole-lyase; the reaction of tryptophan indole-lyase with L-tyrosine resulted in formation of external aldimine, which absorbed at 420 nm, and a very small absorbance peak at 500 nm. 3-Fluoro-L-tyrosine reacted with tryptophan indole-lyase to produce a prominent quinonoid absorbance peak at 500 nm, whereas L-tyrosine, 2-fluoro-L-tyrosine, and all difluoro-L-tyrosines, had a much reduced intensity for this peak. Thus, the presence of ring fluorine substituents in L-tyrosine that are remote from the site of the chemical transformation has significant effects on the rates and equilibria of intermediate formation in the reactions with both tyrosine phenol-lyase and tryptophan indole-lyase. Although it is commonly thought that fluorine substitution will not result in any significant steric effects, our results suggest that the effects of fluorine substitution in the reactions of fluorinated tyrosines with tyrosine phenol-lyase and tryptophan indole-lyase are due to a combination of steric and electronic effects.
通过快速扫描停流分光光度法研究了酪氨酸的环氟化类似物与酪氨酸酚裂解酶和色氨酸吲哚裂解酶（色氨酸酶）的相互作用。L-酪氨酸与酪氨酸酚裂解酶的反应导致在500nm处迅速形成一个小的吸光度峰，这归因于醌类中间体。3-氟-L-酪氨酸与酪氨酸酚裂解酶的反应在500nm处产生一个吸光度更高的峰，3,5-二氟-L-酪氨酸的反应也是如此，这是由于醌类中间体的积累增加。相比之下，与2-氟-L-酪氨酸、2,3-二氟-L-酪氨酸、2,5-二氟-L-酪氨酸和2,6-二氟-L-酪氨酸形成的复合物在500nm处的吸光度强度要低得多。由3-氟-L-酪氨酸形成醌类中间体的速率常数与L-酪氨酸的相当。然而，3,5-二氟-L-酪氨酸反应形成醌类中间体的速率约为L-酪氨酸的一半，而2,3-二氟-L-酪氨酸的反应速率是L-酪氨酸的两倍。此外，2-取代的二氟酪氨酸表现出一种中间体，它迅速形成，在约340nm处有强烈吸收，这可能是由于偕二胺中间体。酪氨酸不是色氨酸吲哚裂解酶的底物；色氨酸吲哚裂解酶与L-酪氨酸的反应导致形成在420nm处有吸收的外部醛亚胺，以及在500nm处有一个非常小的吸光度峰。3-氟-L-酪氨酸与色氨酸吲哚裂解酶反应在500nm处产生一个突出的醌类吸光度峰，而L-酪氨酸、2-氟-L-酪氨酸和所有二氟-L-酪氨酸的这个峰的强度都大大降低。因此，L-酪氨酸中环上远离化学转化位点的氟取代基的存在对与酪氨酸酚裂解酶和色氨酸吲哚裂解酶反应中中间体形成的速率和平衡有显著影响。尽管通常认为氟取代不会产生任何显著的空间效应，但我们的结果表明，氟化酪氨酸与酪氨酸酚裂解酶和色氨酸吲哚裂解酶反应中氟取代的影响是空间效应和电子效应共同作用的结果。
