Abstract:
Thioredoxin reductases (TrxR) activate thioredoxins (Trx) that regulate the activity of diverse target proteins essential to prokaryotic and eukaryotic life. However, very little is understood of TrxR/Trx systems and redox control in methanogenic microbes from the domain Archaea (methanogens), for which genomes are abundant with annotations for ferredoxin:thioredoxin reductases [Fdx/thioredoxin reductase (FTR)] from group 4 of the widespread FTR-like family. Only two from the FTR-like family are characterized: the plant-type FTR from group 1 and FDR from group 6. Herein, the group 4 archetype (AFTR) from Methanosarcina acetivorans was characterized to advance understanding of the family and TrxR/Trx systems in methanogens. The modeled structure of AFTR, together with EPR and Mössbauer spectroscopies, supports a catalytic mechanism similar to plant-type FTR and FDR, albeit with important exceptions. EPR spectroscopy of reduced AFTR identified a transient [4Fe-4S]1+ cluster exhibiting a mixture of S = 7/2 and typical S = 1/2 signals, although rare for proteins containing [4Fe-4S] clusters, it is most likely the on-pathway intermediate in the disulfide reduction. Furthermore, an active site histidine equivalent to residues essential for the activity of plant-type FTR and FDR was found dispensable for AFTR. Finally, a unique thioredoxin system was reconstituted from AFTR, ferredoxin, and Trx2 from M. acetivorans, for which specialized target proteins were identified that are essential for growth and other diverse metabolisms.
摘要:
硫氧还蛋白还原酶(TrxR)激活硫氧还蛋白(Trx),其调节原核生物和真核生物所必需的多种靶蛋白的活性。然而,对来自古菌(产甲烷菌)领域的产甲烷微生物中的TrxR/Trx系统和氧化还原控制知之甚少,其中基因组丰富,具有铁氧还蛋白的注释:来自广泛的FTR样家族第4组的硫氧还蛋白还原酶[Fdx/硫氧还蛋白还原酶(FTR)]。仅表征了来自FTR样家族的两个:来自第1组的植物型FTR和来自第6组的FDR。在这里,来自甲烷杆菌的第4组原型(AFTR)的特征是增进对产甲烷菌家族和TrxR/Trx系统的理解。AFTR的建模结构,连同EPR和Mössbauer光谱,支持类似于植物型FTR和FDR的催化机理,尽管有重要的例外。还原的AFTR的EPR光谱鉴定出瞬态[4Fe-4S]1簇,表现出S=7/2和典型的S=1/2信号的混合,虽然含有[4Fe-4S]簇的蛋白质很少见,它很可能是二硫化物还原中的中间途径。此外,发现与植物型FTR和FDR活性必需的残基等效的活性位点组氨酸对于AFTR是不必要的。最后,从AFTR重建了一个独特的硫氧还蛋白系统,铁氧还蛋白,和来自M.acetivorans的Trx2,为此,确定了对生长和其他多样化代谢至关重要的特定靶蛋白。