Two Biosynthetic Pathways for Aromatic Amino Acids in the Archaeon Methanococcus maripaludis

Abstract

Methanococcus maripaludis is a strictly anaerobic, methane-producing archaeon. Aromatic amino acids (AroAAs) are biosynthesized in this autotroph either by the de novo pathway, with chorismate as an intermediate, or by the incorporation of exogenous aryl acids via indolepyruvate oxidoreductase (IOR). In order to evaluate the roles of these pathways, the gene that encodes the third step in the de novo pathway, 3-dehydroquinate dehydratase (DHQ), was deleted. This mutant required all three AroAAs for growth, and no DHQ activity was detectible in cell extracts, compared to 6.0 ؎ 0.2 mU mg ؊1 in the wild-type extract. The growth requirement for the AroAAs could be fulfilled by the corresponding aryl acids phenylacetate, indoleacetate, and p-hydroxyphenylacetate. The specific incorporation of phenylacetate into phenylalanine by the IOR pathway was demonstrated in vivo by labeling with [1-13 C]phenylacetate. M. maripaludis has two IOR homologs. A deletion mutant for one of these homologs contained 76, 74, and 42% lower activity for phenylpyruvate, p-hydoxyphenylpyruvate, and indolepyruvate oxidation, respectively, than the wild type. Growth of this mutant in minimal medium was inhibited by the aryl acids, but the AroAAs partially restored growth. Genetic complementation of the IOR mutant also restored much of the wild-type phenotype. Thus, aryl acids appear to regulate the expression or activity of the de novo pathway. The aryl acids did not significantly inhibit the activity of the biosynthetic enzymes chorismate mutase, prephenate dehydratase, and prephenate dehydrogenase in cell extracts, so the inhibition of growth was probably not due to an effect on these enzymes.

Disruption of the Operon Encoding Ehb Hydrogenase Limits Anabolic CO 2 Assimilation in the Archaeon Methanococcus maripaludis

“…3. Enzyme systems in M. maripaludis that are expected to require lowpotential electron donors in addition to FMD include CODH-ACS, which catalyzes the biosynthesis of acetyl-CoA from methyltetrahydromethanopterin and CO 2 ; POR, which catalyzes the reductive carboxylation of acetyl-CoA (14, 15); 2-ketoglutarate oxidoreductase, which catalyzes the reductive carboxylation of succinyl-CoA; and branched-chain VOR and two indole-pyruvate oxidoreductases, which are involved in biosynthesis of amino acids from the corresponding carboxylic acids (7,19). In addition, the genome of M. maripaludis contains an oxidoreductase of unknown specificity (9) which could also be coupled to Ehb.…”

Carboxylases in Natural and Synthetic Microbial Pathways

“…In the methanogenic archaea Methanobacterium autotrophicum and Methanococcus maripaludis, novel ferredoxindependent carboxylases were identified that are linked to biosynthetic pathways. Apart from the well-studied pyruvate: ferredoxin oxidoreductase and ␣-ketoglutarate:ferredoxin oxidoreductase, those organisms also employ ␣-keto-isovalerate:ferredoxin oxidoreductase and indolepyruvate:ferredoxin oxidoreductase homologs for the biosynthesis of branched amino acids (e.g., valine) and aromatic amino acids (e.g., tryptophan and phenylalanine) (96,116).…”

Structures of open (R) and close (T) states of prephenate dehydratase (PDT)—Implication of allosteric regulation by l-phenylalanine

“…The enzyme was discovered in 1965 (Cotton and Gibson, 1965) and was initially characterized in bacteria, including several pathogens, as well as in archaea, yeast, fungi and plants ( Fig. 2) (Bode et al, 1984;Fiske and Kane, 1984;Jensen et al, 1988;Porat et al, 2004;Warpeha et al, 2006). The PDT plays a key role in the biosynthesis of L-Phe in organisms that utilize the shikimate pathway (Bentley, 1990;Lingens, 1976).…”