Protein Thiocarboxylate-Dependent Methionine Biosynthesis in <i>Wolinella succinogenes</i>

Krishnamoorthy, Kalyanaraman; Begley, Tadhg P.

doi:10.1021/ja107424t

Cited by 29 publications

(20 citation statements)

References 25 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…OAHS is known to catalyze the conversion of O-acetylhomoserine (OAH) to homocysteine (Hcy) using sulfide directly (Yamagata, 1989). Recent biochemical studies (Krishnamoorthy & Begley, 2011) have suggested that the likely sulfur source in the methionine-biosynthetic pathway of W. succinogenes is a protein thiocarboxylate (Fig. 1).…”

Section: Introductionmentioning

confidence: 99%

A novel mechanism of sulfur transfer catalyzed byO-acetylhomoserine sulfhydrylase in the methionine-biosynthetic pathway ofWolinella succinogenes

Tran

Krishnamoorthy

Begley

et al. 2011

Acta Crystallogr D Biol Cryst

Self Cite

View full text Add to dashboard Cite

O-Acetylhomoserine sulfhydrylase (OAHS) is a pyridoxal 5 0 -phosphate (PLP) dependent sulfide-utilizing enzyme in the l-cysteine and l-methionine biosynthetic pathways of various enteric bacteria and fungi. OAHS catalyzes the conversion of O-acetylhomoserine to homocysteine using sulfide in a process known as direct sulfhydrylation. However, the source of the sulfur has not been identified and no structures of OAHS have been reported in the literature. Here, the crystal structure of Wolinella succinogenes OAHS (MetY) determined at 2.2 Å resolution is reported. MetY crystallized in space group C2 with two monomers in the asymmetric unit. Size-exclusion chromatography, dynamic light scattering and crystal packing indicate that the biological unit is a tetramer in solution. This is further supported by the crystal structure, in which a tetramer is formed using a combination of noncrystallographic and crystallographic twofold axes. A search for structurally homologous proteins revealed that MetY has the same fold as cystathionine -lyase and methionine -lyase. The active sites of these enzymes, which are also PLPdependent, share a high degree of structural similarity, suggesting that MetY belongs to the -elimination subclass of the Cys/Met metabolism PLP-dependent family of enzymes. The structure of MetY, together with biochemical data, provides insight into the mechanism of sulfur transfer to a small molecule via a protein thiocarboxylate intermediate.

show abstract

Section: Introductionmentioning

confidence: 99%

A novel mechanism of sulfur transfer catalyzed byO-acetylhomoserine sulfhydrylase in the methionine-biosynthetic pathway ofWolinella succinogenes

Tran

Krishnamoorthy

Begley

et al. 2011

Acta Crystallogr D Biol Cryst

Self Cite

View full text Add to dashboard Cite

show abstract

“…These two distinct pathways to fosfomycin are an interesting and unusual example of convergent evolution in natural-product biosynthesis. Similar evolutionarily distinct pathways toward the same chemical structures have been described in primary metabolism (e.g., menaquinone, thiamine, lysine, and isopentenyl diphosphate) (10,13,28,32), and a few cases have been described in secondary metabolism (e.g., polyketides) (10).…”

Section: Discussionmentioning

confidence: 63%

Different Biosynthetic Pathways to Fosfomycin in Pseudomonas syringae and Streptomyces Species

Kim

Metcalf

et al. 2012

Antimicrob Agents Chemother

View full text Add to dashboard Cite

cFosfomycin is a wide-spectrum antibiotic that is used clinically to treat acute cystitis in the United States. The compound is produced by several strains of streptomycetes and pseudomonads. We sequenced the biosynthetic gene cluster responsible for fosfomycin production in Pseudomonas syringae PB-5123. Surprisingly, the biosynthetic pathway in this organism is very different from that in Streptomyces fradiae and Streptomyces wedmorensis. The pathways share the first and last steps, involving conversion of phosphoenolpyruvate to phosphonopyruvate (PnPy) and 2-hydroxypropylphosphonate (2-HPP) to fosfomycin, respectively, but the enzymes converting PnPy to 2-HPP are different. The genome of P. syringae PB-5123 lacks a gene encoding the PnPy decarboxylase found in the Streptomyces strains. Instead, it contains a gene coding for a citrate synthase-like enzyme, Psf2, homologous to the proteins that add an acetyl group to PnPy in the biosynthesis of FR-900098 and phosphinothricin. Heterologous expression and purification of Psf2 followed by activity assays confirmed the proposed activity of Psf2. Furthermore, heterologous production of fosfomycin in Pseudomonas aeruginosa from a fosmid encoding the fosfomycin biosynthetic cluster from P. syringae PB-5123 confirmed that the gene cluster is functional. Therefore, two different pathways have evolved to produce this highly potent antimicrobial agent.

show abstract

“…Once genome-wide fitness data from more diverse bacteria is available, we hope to explain many more mysteries. For example, at least one more pathway of homocysteine synthesis remains to be discovered: thermophilic autotrophs from several divisions of bacteria (i.e., Aquifex aeolicus , Pyrolobus fumarii 1A, and Acidimicrobium ferrooxidans DSM 10331) contain neither the traditional nor the DUF39 pathways of homocysteine biosynthesis, nor the protein thiocarboxylate pathway (Krishnamoorthy and Begley 2011) .…”

Section: Discussionmentioning

confidence: 99%

Filling Gaps in Bacterial Amino Acid Biosynthesis Pathways with High-throughput Genetics

Price¹,

Zane²,

Kuehl³

et al. 2017

Preprint

View full text Add to dashboard Cite

For many bacteria with sequenced genomes, we do not understand how they synthesize some amino acids. This makes it challenging to reconstruct their metabolism, and has led to speculation that bacteria might be cross-feeding amino acids. We studied heterotrophic bacteria from 10 different genera that grow without added amino acids even though an automated tool predicts that the bacteria have gaps in their amino acid synthesis pathways. Across these bacteria, there were 11 gaps in their amino acid biosynthesis pathways that we could not fill using current knowledge. Using genome-wide mutant fitness data, we identified novel enzymes that fill 9 of the 11 gaps and hence explain the biosynthesis of methionine, threonine, serine, or histidine by bacteria from six genera. We also found that the sulfate-reducing bacterium Desulfovibrio vulgaris synthesizes homocysteine (which is a precursor to methionine) by using DUF39, NIL/ferredoxin, and COG2122 proteins, and that homoserine is not an intermediate in this pathway. Our results suggest that most free-living bacteria can likely make all 20 amino acids and illustrate how high-throughput genetics can uncover previously-unknown amino acid biosynthesis genes.

show abstract

Protein Thiocarboxylate-Dependent Methionine Biosynthesis in Wolinella succinogenes

Cited by 29 publications

References 25 publications

A novel mechanism of sulfur transfer catalyzed byO-acetylhomoserine sulfhydrylase in the methionine-biosynthetic pathway ofWolinella succinogenes

A novel mechanism of sulfur transfer catalyzed byO-acetylhomoserine sulfhydrylase in the methionine-biosynthetic pathway ofWolinella succinogenes

Different Biosynthetic Pathways to Fosfomycin in Pseudomonas syringae and Streptomyces Species

Filling Gaps in Bacterial Amino Acid Biosynthesis Pathways with High-throughput Genetics

Contact Info

Product

Resources

About