School of Biosciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, UK.
School of Life Sciences, University of Warwick, Coventry, CV4 7AL, UK.
Sci Rep. 2016 Mar 15;6:23191. doi: 10.1038/srep23191.
With the increased incidence of tuberculosis (TB) caused by Mycobacterium tuberculosis there is an urgent need for new and better anti-tubercular drugs. N-succinyl-L,L-diaminopimelic acid desuccinylase (DapE) is a key enzyme in the succinylase pathway for the biosynthesis of meso-diaminopimelic acid (meso-DAP) and L-lysine. DapE is a zinc containing metallohydrolase which hydrolyses N-succinyl L,L diaminopimelic acid (L,L-NSDAP) to L,L-diaminopimelic acid (L,L-DAP) and succinate. M. tuberculosis DapE (MtDapE) was cloned, over-expressed and purified as an N-terminal hexahistidine ((His)6) tagged fusion containing one zinc ion per DapE monomer. We redesigned the DAP synthetic pathway to generate L,L-NSDAP and other L,L-NSDAP derivatives and have characterised MtDapE with these substrates. In contrast to its other Gram negative homologues, the MtDapE was insensitive to inhibition by L-captopril which we show is consistent with novel mycobacterial alterations in the binding site of this drug.
由于结核分枝杆菌引起的结核病发病率不断上升，因此迫切需要新的、更好的抗结核药物。N-琥珀酰-L,L-二氨基庚二酸脱琥珀酰酶（DapE）是合成中隔二氨基庚二酸（meso-DAP）和 L-赖氨酸的琥珀酰酶途径中的关键酶。DapE 是一种含锌的金属水解酶，可将 N-琥珀酰-L,L-二氨基庚二酸（L,L-NSDAP）水解为 L,L-二氨基庚二酸（L,L-DAP）和琥珀酸。结核分枝杆菌 DapE（MtDapE）被克隆、过表达和纯化，作为一个 N 端六组氨酸（(His)6）标记融合蛋白，每个 DapE 单体含有一个锌离子。我们重新设计了 DAP 合成途径，生成 L,L-NSDAP 和其他 L,L-NSDAP 衍生物，并对这些底物进行了 MtDapE 的特性分析。与其他革兰氏阴性同源物不同，MtDapE 对 L-卡普托普利的抑制不敏感，我们的研究表明，这与该药物结合部位新型分枝杆菌的改变一致。
