A periplasmic aldehyde oxidoreductase represents the first molybdopterin cytosine dinucleotide cofactor containing molybdo‐flavoenzyme from Escherichia coli

Abstract

Molybdoenzymes are involved in a large number of enzymatic reactions in the nitrogen, carbon and sulfur cycles. They occur in all the kingdoms of life. With the exception of nitrogenase, all molybdoenzymes carry the molybdenum cofactor (Moco), where the molybdenum atom is coordinated to the unique dithiolene moiety of a conserved tricyclic pyranopterin cofactor called molybdopterin (MPT). Depending on the remaining ligands of the molybdenum center, molybdoenzymes are classified into three families: (a) the xanthine oxidase (XO) family, characterized by a cyanolyzable equatorial sulfur ligand coordinated to the molybdenum atom; (b) the sulfite oxidase family, with two oxo ligands at the molybdenum center; and Three DNA regions carrying genes encoding putative homologs of xanthine dehydrogenases were identified in Escherichia coli, named xdhABC, xdhD, and yagTSRQ. Here, we describe the purification and characterization of gene products of the yagTSRQ operon, a molybdenum-containing ironsulfur flavoprotein from E. coli, which is located in the periplasm. The 135 kDa enzyme comprised a noncovalent (abc) heterotrimer with a large (78.1 kDa) molybdenum cofactor (Moco)-containing YagR subunit, a medium (33.9 kDa) FAD-containing YagS subunit, and a small (21.0 kDa) 2 · [2Fe2S]-containing YagT subunit. YagQ is not a subunit of the mature enzyme, and the protein is expected to be involved in Moco modification and insertion into YagTSR. Analysis of the form of Moco present in YagTSR revealed the presence of the molybdopterin cytosine dinucleotide cofactor. Two different [2Fe2S] clusters, typical for this class of enzyme, were identified by EPR. YagTSR represents the first example of a molybdopterin cytosine dinucleotide-containing protein in E. coli. Kinetic characterization of the enzyme revealed that YagTSR converts a broad spectrum of aldehydes, with a preference for aromatic aldehydes. Ferredoxin instead of NAD + or molecular oxygen was used as terminal electron acceptor. Complete growth inhibition of E. coli cells devoid of genes from the yagTSRQ operon was observed by the addition of cinnamaldehyde to a low-pH medium. This finding shows that YagTSR might have a role in the detoxification of aromatic aldehydes for E. coli under certain growth conditions. Abbreviations ICP-OES, inductively coupled plasma optical emission spectroscopy; MCD, molybdopterin cytosine dinucleotide; MGD, molybdopterin guanine dinucleotide; Moco, molybdenum cofactor; MPT, molybdopterin; Tat, twin arginine protein transport; XDH, xanthine dehydrogenase; XO, xanthine oxidase.

Molecular mechanisms underlying the close association between soil Burkholderia and fungi

“…In addition to these detoxification systems, other proteins with a putative function in detoxification were found to be increased in cocultivations: the proteins YagS (BGLT01120), YagT (BGLT01121) and YagR (BGLT00865), which are part of the yagTSRQ operon, were significantly increased in both co-cultivations (Supplementary Table S3). This operon was shown to encode a periplasmatic aldehyde oxidoreductase that oxidizes a broad spectrum of aldehydes to their respective acids, thereby contributing to the detoxification of the cells (Neumann et al, 2009).…”

Dual RpoH Sigma Factors and Transcriptional Plasticity in a Symbiotic Bacterium

“…Genes with predicted functions include some plausibly involved in stress responses: ecnA, orthologous to an E. coli antitoxin gene whose expression in E coli is induced by osmotic stress in stationary phase (6); orthologs of E. coli yagTSR, a broadspectrum, molybdopterin cytosine dinucleotide cofactor-containing aldehyde dehydrogenase involved in detoxification (44); xseB, predicted to encode a DNA repair enzyme; and SMa1158, which encodes a universal stress protein that is induced in S. meliloti upon exposure to low oxygen and nitric oxide (5,36). Fortythree percent of our rpoH2-dependent genes were induced, and none repressed, during osmotic stress in a previous study (12), suggesting that RpoH2 is important for response to osmotic stress.…”

Identification of a Bis-molybdopterin Intermediate in Molybdenum Cofactor Biosynthesis in Escherichia coli

“…In MCD-containing enzymes, like the periplasmic aldehyde oxidoreductase PaoABC (16), the Moco contains an equatorial sulfido ligand at the molybdenum atom, which is essential for the catalytic activity of this class of enzymes (17). For the final step of MGD biosynthesis, two cofactor molecules are ligated to one molybdenum atom, forming the bis-MGD cofactor (18).…”