Mycothiol Biosynthesis and Metabolic Functions

霉硫醇(Mycothiol，MSH，AcCys-GlcN-ins)是放线菌产生的主要低分子硫醇，是包括链霉菌和分枝杆菌在内的一大类革兰氏阳性细菌。目前的证据表明，MSH在放线菌中具有类似于谷胱甘肽(GSH)的功能，但我们对MSH生物化学的了解仍然有限。本实验室已鉴定了参与MSH生物合成的四种酶和参与MSH依赖的新解毒途径的一种酶。还获得了分枝杆菌突变体，它们在MSH生物合成的各个步骤中被阻断。本研究继续阐明了霉菌硫醇的代谢生化。MSH生物合成的调控将通过MSH生物合成的最后两种酶MSHC和MshD的动力学研究以及通过基因替换研究来测试假定的MSH调节基因的作用。MSH及其生物合成中间体对分枝杆菌对毒素和抗生素敏感性的重要性将通过对MSH缺陷突变株的研究来确定，这些突变株被阻止在MSH生产的各个步骤。通过对污垢分枝杆菌中MSH的放射性标记以鉴定MSH加合物和降解产物，以及通过降解酶霉硫醇S偶联酰胺酶对MSH-利福霉素加合物代谢的动力学研究，将考察MSH在抗生素利福霉素解毒中的作用。已发现一种名为霉硫醇的新型抗氧化剂是由一类细菌产生的，其中包括主要产生抗生素的链霉菌和引起结核病和麻风的分枝杆菌。有证据表明，霉菌硫醇参与保护细胞免受各种毒素和抗生素的伤害。这项研究将阐明分枝杆菌中参与调节分枝杆菌产生分枝杆菌硫醇的因素，以及分枝杆菌硫醇在解毒抗生素利福霉素中的作用。研究结果可为设计提高细菌解毒环境污染物能力的方法(生物修复)和提高链霉菌发酵中抗生素的产量提供依据。

Mycothiol (MSH, AcCys-GlcN-Ins), is the major low-molecular-weight thiol produced by actinomycetes, a broad group of gram positive bacteria which include the streptomycetes and mycobacteria. Current evidence indicates that MSH has functions in actinomycetes analogous to glutathione (GSH) in GSH-producing organisms but our understanding of MSH biochemistry is still limited. The four enzymes involved in MSH biosynthesis and one enzyme involved in a novel MSH-dependent detoxification pathway have been identified in this laboratory. Mycobacterial mutants have also been obtained which are blocked at various steps of MSH biosynthesis. The present studies continue the elucidation of the metabolic biochemistry of mycothiol. The regulation of MSH biosynthesis will be examined through kinetic studies with the final two enzymes of MSH biosynthesis, MshC and MshD, and through gene replacement studies to test the role of putative MSH regulatory genes. The importance of MSH and its biosynthetic intermediates for mycobacterial sensitivity to toxins and antibiotics will be determined through studies of MSH deficient mutants blocked at various steps of MSH production. The role of MSH in the detoxification of the antibiotic rifamycin will be examined using radioactive labeling of MSH in Mycobacterium smegmatis to permit identification of MSH adducts and degradation products, and through kinetic studies of MSH-rifamycin adduct metabolism by the degradative enzyme mycothiol S-conjugate amidase.A novel antioxidant named mycothiol has been found to be produced by a class of bacteria which include the streptomycetes, the main antibiotic-producing bacteria, and the mycobacteria, the causative agents of tuberculosis and leprosy. Evidence indicates that mycothiol is involved in protecting cells against various toxins and antibiotics. This research will elucidate the factors involved in regulating mycothiol production in mycobacteria and the role of mycothiol in detoxifying the antibiotic rifamycin. The results could provide the basis for devising methods to enhance the ability of bacteria to detoxify environmental pollutants (bioremediation) and for increasing antibiotic production in streptomycete fermentations.