Mycothiol Biosynthesis and Metabolic Functions

真菌硫醇（MSH, AcCys-GlcN-Ins）是放线菌产生的主要低分子量硫醇，放线菌是一大类革兰氏阳性细菌，包括链菌和分枝杆菌。目前的证据表明，MSH在放线菌中具有类似于GSH产生生物体内谷胱甘肽（GSH）的功能，但我们对MSH生物化学的理解仍然有限。参与MSH生物合成的四种酶和参与一种新的MSH依赖性解毒途径的一种酶已经在这个实验室被确定。分枝杆菌突变体也在MSH生物合成的各个步骤中被阻断。目前的研究继续阐明菌硫醇的代谢生化。通过MSH生物合成的最后两种酶MshC和MshD的动力学研究，以及通过基因替代研究来测试假定的MSH调节基因的作用，将对MSH生物合成的调节进行研究。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研究在MSH生产的各个步骤中被阻断的MSH缺陷突变体来确定。MSH在抗生素利福霉素解毒中的作用将通过对耻毛分枝杆菌中MSH的放射性标记来检测MSH加合物和降解产物，并通过降解酶真菌硫醇s偶联氨基酶对MSH-利福霉素加合物代谢的动力学研究。一种新型抗氧化剂——真菌硫醇已经被发现是由一类细菌产生的，这些细菌包括链霉菌，主要的抗生素产生细菌，以及分枝杆菌，结核病和麻风病的病原体。有证据表明，真菌硫醇参与保护细胞免受各种毒素和抗生素的侵害。本研究将阐明分枝杆菌中调节菌硫醇产生的因素以及菌硫醇在利福霉素解毒中的作用。该结果可为设计提高细菌对环境污染物解毒能力（生物修复）和提高链霉菌发酵过程中抗生素产量的方法提供依据。

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.