Metabolic Pathway Rerouting in Paraburkholderia rhizoxinica Evolved Long-Overlooked Derivatives of Coenzyme F₄₂₀

Abstract

Author contributions and conflict of interest Daniel Braga performed research, analyzed data (molecular biology, malachite green assay, mass spectrometry) and contributed to writing the manuscript, Daniel Last performed research and analyzed data (structure elucidation, Fno assay, biogas plant studies), Mahmudul Hasan performed research (CofC/D enzyme assays), Huijuan Guo performed research and analyzed data (structure elucidation), Daniel Leichnitz performed research (chemical synthesis), Zerrin Uzum performed research (microscopy), Ingrid Richter performed research (microscopy), Felix Schalk performed research (cofE constructs), Christine Beemelmanns designed research, acquired funding, analyzed data (structure elucidation, synthesis) and edited the manuscript, Christian Hertweck designed research, acquired funding and edited the manuscript, Gerald Lackner designed the study, acquired funding and wrote the original manuscript. The authors declare no conflict of interest.

Cellular and Structural Basis of Synthesis of the Unique Intermediate Dehydro-F 420 -0 in Mycobacteria

“…Integrating these findings with other recent literature, it is now clear that the substrate for the initial stage of F 420 tail biosynthesis differs between F 420 -producing organisms (19,24,26). We definitively show here that, in mycobacteria, PEP is the substrate for F 420 biosynthesis, resolving the ambiguity in the literature (24,26).…”

“…The study also showed that CofC from Methanocaldococcus jannaschii utilized PEP rather than 2PL for F 420 biosynthesis (24), leading the authors to conclude that PEP is the general precursor for F 420 biosynthesis in microorganisms. However, these findings contradict previous analysis of CofC activity in M. jannaschii cell lysates (19,21), as well as recent biochemical analysis, which shows that CofC preferentially utilizes 2PL for F 420 biosynthesis (26). In turn, these findings cast doubt on whether PEP is truly the preferred substrate for mycobacterial F 420 biosynthesis and whether DH-F 420 -0 is the physiological intermediate in this pathway.…”

“…We definitively show here that, in mycobacteria, PEP is the substrate for F 420 biosynthesis, resolving the ambiguity in the literature (24,26). In contrast, in the archaeal and proteobacterial species that have been analyzed, 2PL and 3PG, respectively, are preferentially utilized (19,26). This divergent substrate utilization occurs despite the enzymes responsible for this stage of synthesis (FbiD/CofC and FbiA/CofD) sharing a common evolutionary history (8).…”

“…This suggests that the substrate specificity of these enzymes has evolved in response to selection to maintain compatibility between the substrate used for F 420 biosynthesis and what is available in the cellular metabolite pool. Both PEP and 3PG are intermediates in central metabolic pathways, including glycolysis, whereas 2PL is not thought to be present in significant quantities in most organisms (24,26,32). This makes PEP and 3PG compatible substrates for F 420 biosynthesis in Mycobacterium spp.…”

Cofactor F420-Dependent Enzymes: An Under-Explored Resource for Asymmetric Redox Biocatalysis

“…In addition to the engineering considerations that this may impose, it may also be responsible for the biochemical diversity of this step in different microorganisms. In different microbes, the CofC/FbiD-dependent step uses 2-phospholactate [75], 3-phosphoglycerate [76] or phosphoenolpyruvate [59] as a substrate, which may reflect the relative abundance of those metabolites in various bacteria and archaea and the thermodynamic constraints on this step.…”