Department of Biochemistry and Molecular Pharmacology, UMass Medical School, 364 Plantation Street, Worcester, Massachusetts 01605, United States.
Program in Chemical Biology, UMass Medical School, 364 Plantation Street, Worcester, Massachusetts 01605, United States.
Biochemistry. 2020 Mar 3;59(8):933-942. doi: 10.1021/acs.biochem.9b01078. Epub 2020 Feb 17.
Sterile alpha and toll/interleukin receptor (TIR) motif-containing protein 1 (SARM1) plays a pivotal role in triggering the neurodegenerative processes that underlie peripheral neuropathies, traumatic brain injury, and neurodegenerative diseases. Importantly, SARM1 knockdown or knockout prevents degeneration, thereby demonstrating that SARM1 is a promising therapeutic target. Recently, SARM1 was shown to promote neurodegeneration via its ability to hydrolyze NAD, forming nicotinamide and ADP ribose (ADPR). Herein, we describe the initial kinetic characterization of full-length SARM1, as well as the truncated constructs corresponding to the SAMTIR and TIR domains, highlighting the distinct challenges that have complicated efforts to characterize this enzyme. Moreover, we show that bacterially expressed full-length SARM1 (/ = 6000 ± 2000 M s) is at least as active as the TIR domain alone (/ = 1500 ± 300 M s). Finally, we show that the SARM1 hydrolyzes NAD via an ordered uni-bi reaction in which nicotinamide is released prior to ADPR.
无菌α和 Toll/白细胞介素受体（TIR）结构域包含蛋白 1（SARM1）在触发外周神经病变、创伤性脑损伤和神经退行性疾病的神经退行性过程中起着关键作用。重要的是，SARM1 的敲低或敲除可防止退化，从而表明 SARM1 是一个有前途的治疗靶点。最近，SARM1 被证明通过其水解 NAD 的能力促进神经退行性变，形成烟酰胺和 ADP 核糖（ADPR）。在此，我们描述了全长 SARM1 的初始动力学特征，以及对应于 SAMTIR 和 TIR 结构域的截断构建体，突出了使该酶的特征复杂化的独特挑战。此外，我们表明细菌表达的全长 SARM1（/ = 6000 ± 2000 M s）的活性至少与单独的 TIR 结构域一样高（/ = 1500 ± 300 M s）。最后，我们表明 SARM1 通过顺序的单双反应水解 NAD，其中烟酰胺在 ADPR 之前释放。
