Maity Amarendra Nath, Chen Jun-Ru, Ke Ting-Xi, Ke Shyue-Chu
Department of Physics, National Dong Hwa University, Hualien 97401, Taiwan.
Department of Occupational Safety and Health, Chang Jung Christian University, Tainan 711301, Taiwan.
Int J Mol Sci. 2025 Sep 3;26(17):8561. doi: 10.3390/ijms26178561.
The ability of enzymes to access various conformational states is often essential for their catalytic activity. Lysine 5,6-aminomutase (5,6-LAM), a pyridoxal 5'-phosphate (PLP) and 5'-deoxyadenosylcobalamin (dAdoCbl)dependent enzyme, catalyzes 1,2-amino shift in lysine isomers by shuttling between an open conformational state and a closed conformational state. Nevertheless, suicide inactivation of 5,6-LAM is an obstacle to the realization of its potential as a biocatalyst. In this work, the fate of the reaction of 5-hydroxylysine, an analogue of lysine, is investigated using spectroscopic and computational methods. Although 5-hydroxylysine does not afford any product, results obtained from UV-visible and electron paramagnetic resonance (EPR) spectroscopies demonstrate that initial steps of the catalytic cycle are performed with it. Simulation of the weakly spin-coupled spectrum estimates an intermediate distance between the PLP substrate-based radical and Co(II) in comparison to the that in the open state and the closed state. This distinct conformational state, different from the open state and the closed state, is alluded to in its putative role in suicide inactivation and denoted as the suicidally-inactivated state. Our findings highlight the emergence of EPR spectroscopy as a powerful tool to uncover the hidden conformations in radical enzymes. These results provide new insights into the suicide inactivation of dAdoCbl-dependent enzymes.
酶进入各种构象状态的能力通常对其催化活性至关重要。赖氨酸5,6-氨基变位酶（5,6-LAM）是一种依赖于磷酸吡哆醛（PLP）和5'-脱氧腺苷钴胺素（dAdoCbl）的酶，通过在开放构象状态和封闭构象状态之间穿梭来催化赖氨酸异构体中的1,2-氨基转移。然而，5,6-LAM的自杀失活是阻碍其作为生物催化剂发挥潜力的一个障碍。在这项工作中，使用光谱学和计算方法研究了赖氨酸类似物5-羟基赖氨酸的反应命运。尽管5-羟基赖氨酸没有产生任何产物，但紫外可见光谱和电子顺磁共振（EPR）光谱获得的结果表明，催化循环的初始步骤是与之进行的。与开放状态和封闭状态相比，对弱自旋耦合光谱的模拟估计了基于PLP底物的自由基与Co（II）之间的中间距离。这种不同于开放状态和封闭状态的独特构象状态，在其自杀失活的假定作用中有所提及，并被称为自杀失活状态。我们的研究结果突出了EPR光谱作为揭示自由基酶中隐藏构象的强大工具的出现。这些结果为依赖dAdoCbl的酶的自杀失活提供了新的见解。
