Purification, Characterization and Inhibition by Fluoride of Enolase from Streptococcus mutans DSM320523
Purification, Characterization and Inhibition by Fluoride of Enolase from Streptococcus mutans DSM320523
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变形链球菌烯醇酶的纯化、表征和氟化物抑制 DSM320523
DOI：
10.1159/000261494
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发表时间：
1992
期刊：
影响因子：
4.2
通讯作者：
P. Bartholmes
中科院分区：
文献类型：
--
作者：
M. Kaufmann;P. Bartholmes
变形链球菌烯醇化酶经三步纯化得到纯品。如所示的分析超离心和聚丙烯酰胺凝胶电泳在变性条件下，纯化的酶是一个八聚体的分子量Mr = 360 kDa，由8个相同的亚基与Mr = 45 kDa。测定了S.在1 mM Mg ~（2+）存在下，变形杆菌烯醇化酶的活性为Aspec = 130 IU × mg ~（-1），KM = 0.44 mM。酶活性被0 - 10 mM NaF范围内的氟化物非竞争性抑制，产生Ki = Ki = 1.39 mM，但当测试浓度高于10 mM时，抑制特性变得具有竞争性。（0.5 mM）氟化物的抑制作用显著增强，产生Ki = 0.26 mM。NaPO 3F的抑制与Ki = 0.9 mM竞争，这表明游离氟离子与磷酸盐组合在抑制S.变形烯醇化酶。由此可见，氟和磷联合抑制烯醇化酶可影响糖酵解。变形杆菌等降低酸的产生或甚至生长速率，从而导致潜在的防龋作用。
Enolase from Streptococcus mutans has been purified to homogeneity by a three-step procedure. As shown by analytical ultracentrifugation and poly-acrylamide gel electrophoresis under denaturing conditions, the purified enzyme is an octamer with molecular weight Mr = 360 kDa, composed of eight identical subunits with Mr = 45 kDa. The kinetic parameters of S. mutans enolase in the presence of 1 mM Mg2+ are Aspec = 130IU × mg-1 and KM = 0.44 mM as determined by steady-state experiments in the D-glycerate 2-phosphate to enolpyruvate phosphate reaction. Enzymatic activity is inhibited noncompetitively by fluoride in the range between 0 and 10 mM NaF yielding Ki = Ki = 1.39 mM, but inhibition characteristics become competitive when tested above 10 mM. In the presence of only small amounts of phosphate (0.5 mM) the inhibitory effect of fluoride is enhanced dramatically yielding Ki = 0.26 mM. Inhibition by NaPO3F is competitive with Ki = 0.9 mM, indicating that free fluoride ions in combination with phosphate are more effective in inhibiting S. mutans enolase. It can be concluded that inhibition of enolase by fluoride in combination with phosphate can influence glycolysis in S. mutans and so reduce acid production or even growth rate, thereby leading to potential anticariogenic effects.