Methanococci Use the Diaminopimelate Aminotransferase (DapL) Pathway for Lysine Biosynthesis

Abstract

The pathway of lysine biosynthesis in the methanococci has not been identified previously. A variant of the diaminopimelic acid (DAP) pathway uses diaminopimelate aminotransferase (DapL) to catalyze the direct conversion of tetrahydrodipicolinate (THDPA) to LL-DAP. Recently, the enzyme DapL (MTH52) was identified in Methanothermobacter thermautotrophicus and shown to belong to the DapL1 group. Although the Methanococcus maripaludis genome lacks a gene that can be unambiguously assigned a DapL function based on sequence similarity, the open reading frame MMP1527 product shares 30% amino acid sequence identity with MTH52. A ⌬mmp1527 deletion mutant was constructed and found to be a lysine auxotroph, suggesting that this DapL homolog in methanococci is required for lysine biosynthesis. In cell extracts of the M. maripaludis wild-type strain, the specific activity of DapL using LL-DAP and ␣-ketoglutarate as substrates was 24.3 ؎ 2.0 nmol min ؊1 mg of protein ؊1 . The gene encoding the DapL homolog in Methanocaldococcus jannaschii (MJ1391) was cloned and expressed in Escherichia coli, and the protein was purified. The maximum activity of MJ1391 was observed at 70°C and pH 8.0 to 9.0. The apparent K m s of MJ1391 for LL-DAP and ␣-ketoglutarate were 82.8 ؎ 10 M and 0.42 ؎ 0.02 mM, respectively. MJ1391 was not able to use succinyl-DAP or acetyl-DAP as a substrate. Phylogenetic analyses suggested that two lateral gene transfers occurred in the DapL genes, one from the archaea to the bacteria in the DapL2 group and one from the bacteria to the archaea in the DapL1 group. These results demonstrated that the DapL pathway is present in marine methanogens belonging to the Methanococcales.

Biochemical Characterization of UDP-N-acetylmuramoyl-L-alanyl-D-glutamate: meso-2,6-diaminopimelate ligase (MurE) from Verrucomicrobium spinosum DSM 4136T

“…V. spinosum was found to employ the recently discovered l , l -diaminopimelate aminotransferase (DapL) pathway [6], [7], [8], [9] as the sole route for the synthesis of diaminopimelate (A 2 pm) and l -lysine ( l -Lys), based on biochemical and bioinformatical evidence [10]. In the anabolism of PG, the penultimate intermediate in the l -lysine biosynthesis pathway, meso -diaminopimelate ( meso -A 2 pm), serves as one of the cross-linking amino acids in Gram-negative bacteria, and l -Lys serves the same purpose in many Gram-positive bacteria [11].…”

Structure and Function of Cyanobacterial DHDPS and DHDPR

“…1A)11011. The pathway then diverges into four sub-pathways, namely the acetylase, aminotransferase, dehydrogenase and succinylase pathways, which operate across different genera and kingdoms21213. For example, the aminotransferase pathway is canonical to plants, but is also innate to cyanobacteria, including Anabaena variabilis (Kyoto Encyclopedia of Genes and Genomes www.genome.jp/kegg-bin/show_module?ava_M00527+Ava_3607, 2016).…”

Enzymology of Bacterial Lysine Biosynthesis

“…This transamination reaction utilises glutamate as an amino donor to yield -ketoglutarate. (Hudson et al, , 2008Liu et al, 2010;McCoy et al, 2006) The enzyme was first isolated from plant and cyanobacterial species and thus demonstrated a new branch of the lysine biosynthesis pathway existed . Although plants are known to synthesise lysine de novo, components of the pathway required for conversion of THDP to meso-DAP had not been identified previously despite years of investigation.…”

“…More recently LL-DAP-AT has been identified in algal, archaeal and bacterial species including, Chlamydia trachomatis (McCoy et al, 2006), Chlamydomonas reihardtii , Methanocaldococcus jannaschii (Liu et al, 2010), and Protochlamydia amoebophila (McCoy et al, 2006). Comparative genomic analyses shows that LL-DAP-AT is restricted to the eubacterial lineages, Bacteroidetes, Chlamydiae, Chloroflexi, Cyanobacteria, Desulfuromonadales, Firmicutes, and Spirochaeta; and the archaea, Archaeoglobaceae and Methanobacteriaceae (Hudson et al, 2008).…”