Tsakiris Stylianos, Schulpis Kleopatra H
Department of Experimental Physiology, Medical School, University of Athens, Greece.
Z Naturforsch C J Biosci. 2002 May-Jun;57(5-6):506-11. doi: 10.1515/znc-2002-5-618.
The aim of this work was to evaluate, in vitro, the effect of L-alanine (Ala) on suckling rat brain acetylcholinesterase (AChE) and on eel Electrophorus electricus pure AChE inhibited by L-phenylalanine (Phe) as well as to investigate whether Phe or Ala is a competitive inhibitor or an effector of the enzyme. AChE activity was determined in brain homogenates and in the pure enzyme after 1 h preincubation with 1.2 mM of Phe or Ala as well as with Phe plus Ala. The activity of the pure AChE was also determined using as a substrate different amounts of acetylthiocholine. Ala reversed completely the inhibited AChE by Phe (18-20% in 500-600 microM substrate, p<0.01). Lineweaver-Burk plots showed that Vmax remained unchanged. However, Km was found increased with Phe (150%, p<0.001), decreased with Ala alone (50%, p<0.001) and unaltered with Phe plus Ala. It is suggested that: a) Phe presents a competitive inhibitory action with the substrate whereas Ala a competitive activation; b) Ala competition with Phe might unbind the latter from AChE molecule inducing the enzyme stimulation; c) Ala might reverse the inhibitory effect of Phe on brain AChE in phenylketonuric patients, if these results are extended into the in vivo reality.
本研究旨在体外评估L-丙氨酸（Ala）对乳鼠脑乙酰胆碱酯酶（AChE）以及对被L-苯丙氨酸（Phe）抑制的电鳗纯AChE的影响，并研究Phe或Ala是该酶的竞争性抑制剂还是效应物。在与1.2 mM的Phe或Ala以及Phe加Ala预孵育1小时后，测定脑匀浆和纯酶中的AChE活性。还使用不同量的乙酰硫代胆碱作为底物测定纯AChE的活性。Ala完全逆转了Phe对AChE的抑制作用（在500 - 600 microM底物中为18 - 20%，p<0.01）。Lineweaver - Burk图显示Vmax保持不变。然而，发现Km随Phe增加（150%，p<0.001），单独使用Ala时降低（50%，p<0.001），Phe加Ala时不变。提示：a）Phe对底物呈现竞争性抑制作用，而Ala呈现竞争性激活作用；b）Ala与Phe竞争可能使Phe从AChE分子上解离，从而诱导酶的激活；c）如果这些结果能扩展到体内情况，Ala可能逆转苯丙酮尿症患者中Phe对脑AChE的抑制作用。
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