Biology subjects

Daumann, L. J.

Publications and source records attributed to Daumann, L. J..

2 recordsLinked to original sources

bioRxiv2023

Assessing Lanthanide-Dependent Methanol Dehydrogenase Activity: The Assay Matters

Artificial dye-coupled assays have been widely adopted as a rapid and convenient method to assess the activity of methanol dehydrogenases (MDH). Lanthanide(Ln)-dependent XoxF-MDHs from methanotrophs and methylotrophs are able to incorporate different lanthanides (Lns) in their active site. The artificial dye-coupled assay showed that the earlier Lns exhibit a higher enzyme activity than the late Lns. Although this assay is widely used, there are limitations. It is not unusual that a pH of 9 is required and that activators like ammonium have to be added to the assay mixture which do not reflect the conditions inside the cell. Moreover, different Ln-MDH variants are not obtained by the direct isolation from the cells grown with the respective Ln, but by metal titration of the Ln in a partial-apo-MDH or by incubation of an apo-MDH with the Ln. Herein, we report the cultivation of Ln-dependent methanotroph Methylacidiphilum fumariolicum SolV with nine different Lns, the isolation of the respective MDH and the assessment of the enzyme activity using the artificial dye-coupled assay. We compare these results with an adapted protein-coupled assay using the physiological partner and electron acceptor cytochrome cGJ (cyt cGJ) instead of artificial dyes. We demonstrate that, depending on the assay, two distinct trends are observed among the Ln series. The specific activity of La-, Ce- and Pr-MDH, as measured by the protein-coupled assay, exceeds the specific enzyme activity measured by the dye-coupled assay. This suggests that the early Lns also have a positive effect on the interaction between XoxF-MDH and its cyt cGJ thereby increasing functional efficiency.

biochemistry↗

bioRxiv2022

Identification of a biosynthetic gene cluster encoding a novel lanthanide chelator in Methylorubrum extorquens AM1

Many bacteria secrete metallophores, low-molecular weight organic compounds that bind ions with high selectivity and affinity, in order to access essential metals from the environment.1 Previous work has elucidated the structures and biosynthetic machinery of metallophores specific for iron, zinc, nickel, molybdenum, and copper.1 No lanthanide-specific metallophore has been discovered despite the knowledge that lanthanide metals (Ln) have been revealed to be essential cofactors for certain alcohol dehydrogenases across a diverse range of phyla.2 Here, we report the biosynthetic machinery, the structure, and the physiological relevance of the first known lanthanophore, methylolanthanin. The structure of methylolanthanin exhibits a unique 4-hydroxybenzoate moiety which has not previously been described in other metallophores. We find that production of methylolanthanin is required for normal levels of Ln accumulation in the methylotrophic bacterium Methylobacterium extorquens AM1, while overexpression of the molecule greatly increases bioaccumulation. Our results provide a clearer understanding of how Ln-utilizing bacteria sense, scavenge, and store Ln; essential processes in the environment where Ln are poorly bioavailable. Beyond Ln, we anticipate our study to be a starting point for understanding how organisms acquire other f-block metals, the actinides.3 More broadly, the discovery of a lanthanophore opens doors for study of how biosynthetic gene clusters are repurposed for new functions, how metallophores acquire their metal specificity, and the complex relationship between metal homeostasis and fitness.

microbiology↗