Human platelet 12-lipoxygenase: Naturally occurring Q261/R261 variants and N544L mutant show altered activity but unaffected substrate binding and membrane association behavior
Human platelet 12-lipoxygenase: Naturally occurring Q261/R261 variants and N544L mutant show altered activity but unaffected substrate binding and membrane association behavior
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DOI：
10.3892/ijmm_00000289
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发表时间：
2009-12-01
影响因子：
5.4
通讯作者：
Skrzypczak-Jankun, Ewa
中科院分区：
文献类型：
--
作者：
Aleem, Ansari M.;Wells, Leigh;Skrzypczak-Jankun, Ewa
人类血小板12-脂氧合酶的单核苷酸多态性（SNP）R261 Q与多种人类疾病相关。为了更好地理解生物学性能，我们比较了重组酶的酶性质：“野生型”，如在酶外周具有单个Q261 R突变的Q261和R261变体，以及具有改变的活性位点的N544 L突变体。R261变体不遵循相同的动力学，例如WT-Q261，其显示滞后期，产物的较慢积累，遵循不同的时间过程而没有达到Q261变体的平台特征。活性位点中的N544 L取代几乎根除了酶活性，证明天冬酰胺对于催化作用与保守的组氨酸和C-末端异亮氨酸一样重要。所有三种酶具有相当的底物结合和膜缔合行为。我们的结论是，天然存在的SNP，在远离活性位点的位置引起单突变，可以改变这种寡聚酶的蛋白质-蛋白质缔合，从而影响变构机制的动力学特性和在亚膜前沿的分子识别/信号传导。
The single nucleotide polymorphism (SNP) R261Q in the human platelet 12-lipoxygenase has been correlated with several human diseases. To understand better the biological performance we have compared enzymatic properties of the recombinant enzymes: 'wild-type' as Q261 and R261 variants with a single Q261R mutation at the enzyme periphery and N544L mutant with an altered active site. The R261 variant does not follow the same kinetics such as WT-Q261 showing a lag phase, a slower accumulation of product, following a different time-course without reaching plateau characteristic for the Q261 variant. The N544L substitution in the active site almost eradicates enzymatic activity proving that asparagine is as important for catalysis as the conserved histidines and C-terminal isoleucine. All three enzymes have comparable substrate binding and membrane association behavior. We conclude that the naturally occurring SNP, causing single mutation at a location distant to the active site, can alter the protein-protein association of this oligomeric enzyme making impact on kinetic properties of an allosteric mechanism and molecular recognition/signaling at a submembrane frontier.