Thioredoxin targets fundamental processes in a methane-producing archaeon, Methanocaldococcus jannaschii

Abstract

Thioredoxin (Trx), a small redox protein, controls multiple processes in eukaryotes and bacteria by changing the thiol redox status of selected proteins. The function of Trx in archaea is, however, unexplored. To help fill this gap, we have investigated this aspect in methanarchaea-strict anaerobes that produce methane, a fuel and greenhouse gas. Bioinformatic analyses suggested that Trx is nearly universal in methanogens. Ancient methanogens that produce methane almost exclusively from H 2 plus CO 2 carried approximately two Trx homologs, whereas nutritionally versatile members possessed four to eight. Due to its simplicity, we studied the Trx system of Methanocaldococcus jannaschii-a deeply rooted hyperthermophilic methanogen growing only on H 2 plus CO 2 . The organism carried two Trx homologs, canonical Trx1 that reduced insulin and accepted electrons from Escherichia coli thioredoxin reductase and atypical Trx2. Proteomic analyses with air-oxidized extracts treated with reduced Trx1 revealed 152 potential targets representing a range of processes-including methanogenesis, biosynthesis, transcription, translation, and oxidative response. In enzyme assays, Trx1 activated two selected targets following partial deactivation by O 2 , validating proteomics observations: methylenetetrahydromethanopterin dehydrogenase, a methanogenesis enzyme, and sulfite reductase, a detoxification enzyme. The results suggest that Trx assists methanogens in combating oxidative stress and synchronizing metabolic activities with availability of reductant, making it a critical factor in the global carbon cycle and methane emission. Because methanogenesis developed before the oxygenation of Earth, it seems possible that Trx functioned originally in metabolic regulation independently of O 2 , thus raising the question whether a complex biological system of this type evolved at least 2.5 billion years ago. methanogenic archaea | redox regulation | hydrothermal vent | early Earth | evolution T hioredoxins (Trxs) are small (∼12-kDa) redox proteins typically bearing a characteristic Cys-Gly-Pro-Cys motif that reduce specific disulfide bonds of selected proteins (1). Reduction alters the biochemical properties of the proteins targeted-e.g., by increasing their activity or solubility (1). Trxs are found in the three domains of life: bacteria, eukarya, and archaea (2). In eukarya and bacteria, the regulatory role of Trx has been shown to span the major aspects of metabolism, including photosynthesis, biosynthesis, replication, transcription, translation, and stress response (1). Trx also acts as an electron donor for enzymes, notably ribonucleotide reductase, phosphoadenosinephosphosulfate reductase, methionine sulfoxide reductase, and peroxiredoxins (1). However, in contrast to the wealth of information for bacteria and eukaryotes, our understanding of archaeal Trx is limited to its biochemical and structural properties (3-9). Its physiological role remains a mystery.To help fill this gap, we have investigated the role of Trx in a group of a...

A Novel F420-dependent Thioredoxin Reductase Gated by Low Potential FAD

“…Similarly, the properties and distribution of DFTR and the conditions prevailing in M. jannaschii's habitat lead to the proposal that this enzyme developed in the low redox environment of early Earth and gave rise to NTR. This implication fits the hypothesis that the Trx-based redox control developed on Earth long before oxygen appeared (20).…”

“…Similarly, there have been several reports on the Trx and TrxRs of anaerobic archaea (9,15,(17)(18)(19)(20)(21)(22)(23), including the methanogenic archaea, which are strict anaerobes (9,17,18,(21)(22)(23). Recently, a proteomics study has shown that there is clear potential for a Trx-based global redox regulation of metabolism, including methanogenesis, in Methanocaldococcus jannaschii, a strictly anaerobic and deeply rooted hyperthermophilic methanogenic archaeon that lives in deep-sea hydrothermal vents (20). Two reports also describe the activation of deactivated methanogen proteins by thioredoxin (10,20).…”

“…This archaeon carries two Trxs, Mj-Trx1 which is a typical Trx, and Mj-Trx2 with unknown activity, and a TrxR (Mj-TrxR) that is an NTR homolog yet does not use NAD(P)H (20). We demonstrate that Mj-TrxR uses coenzyme F 420 , a deazaflavin derivative present in all methanogens, as electron carrier.…”

“…However, it has also been investigated significantly in the anaerobic organisms (6 -24) focusing on both the characteristics of the Trxs and TrxRs (6,8,9,(11)(12)(13)(15)(16)(17)(18)(19)(20)(21)(22)(23) and their physiological roles (6,7,10,14,20). For example, the isolation of the putative Trx targets via Trxaffinity chromatography and the results from activation assays with select purified and deactivated enzymes have established that in Chlorobaculum tepidum, a green sulfur bacterium and an anaerobic phototroph, the thioredoxin system implements post-translational control on the tricarboxylic acid or the TCA cycle and sulfur metabolism and assists in the defense against oxidative stress (7).…”

“…Recently, a proteomics study has shown that there is clear potential for a Trx-based global redox regulation of metabolism, including methanogenesis, in Methanocaldococcus jannaschii, a strictly anaerobic and deeply rooted hyperthermophilic methanogenic archaeon that lives in deep-sea hydrothermal vents (20). Two reports also describe the activation of deactivated methanogen proteins by thioredoxin (10,20).…”