The cellular response towards lanthanum is substrate specific and reveals a novel route for glycerol metabolism inPseudomonas putidaKT2440

Abstract

<jats:title>Abstract</jats:title><jats:p>Ever since the discovery of the first rare earth element (REE)-dependent enzyme, the physiological role of lanthanides has become an emerging field of research due to the potential environmental implications and biotechnological opportunities. In <jats:italic>Pseudomonas putida</jats:italic> KT2440, the two pyrroloquinoline quinone-dependent alcohol dehydrogenases (PQQ-ADHs) PedE and PedH are inversely produced in response to La<jats:sup>3+</jats:sup>-availability. This REE-switch is orchestrated by a complex regulatory network including the PedR2/PedS2 two-component system and is important for efficient growth on several alcoholic volatiles. As <jats:italic>P. putida</jats:italic> is exposed to a broad variety of organic compounds in its natural soil habitat, the cellular responses towards La<jats:sup>3+</jats:sup> during growth on various carbon and energy sources were investigated with a differential proteomic approach. Apart from the Ca<jats:sup>2+</jats:sup>-dependent enzyme PedE, the differential abundance of most other identified proteins was conditional and revealed a substrate specificity. Concomitant with the proteomic changes, La<jats:sup>3+</jats:sup> had a beneficial effect on lag-phases while causing reduced growth rates and lower optical densities in stationary phase during growth on glycerol. When these growth phenotypes were evaluated with mutant strains, a novel metabolic route for glycerol utilization was identified that seems to be functional in parallel with the main degradation pathway encoded by the <jats:italic>glpFKRD</jats:italic> operon. The newly discovered route is initiated by PedE and/or PedH, which most likely convert glycerol to glyceraldehyde. In the presence of lanthanum, glyceraldehyde seems to be further oxidized to glycerate, which, upon phosphorylation to glycerate-2-phosphate by the glycerate kinase GarK, is finally channelled into the central metabolism.</jats:p><jats:sec><jats:title>Importance</jats:title><jats:p>The biological role of rare earth elements has long been underestimated and research has mainly focused on methanotrophic bacteria. We have recently demonstrated that <jats:italic>P. putida,</jats:italic> a plant growth promoting bacterium that thrives in the rhizosphere of various feed crops, possesses a REE-dependent alcohol dehydrogenase (PedH), but knowledge about lanthanide-dependent effects on physiological traits in non-methylotrophic bacteria is still scarce. This study demonstrates that the cellular response of <jats:italic>P. putida</jats:italic> KT2440 towards La<jats:sup>3+</jats:sup> is mostly substrate specific and that during growth on glycerol, La<jats:sup>3+</jats:sup> has a severe effect on several growth parameters. We provide compelling evidence that the observed physiological changes are linked to the catalytic activity of PedH and thereby identify a novel route for glycerol metabolism in this biotechnological relevant organism. Overall, these findings demonstrate that lanthanides can alter important physiological traits of non-methylotrophic bacteria, which might consequently influence their competitiveness during colonization of various environmental niches.</jats:p></jats:sec>

Rare Earth Element (REE)-Dependent Growth of Pseudomonas putida KT2440 Relies on the ABC-Transporter PedA1A2BC and Is Influenced by Iron Availability

“…It is, however, possible that additional genes/proteins required for PedH-dependent growth rely on, or are regulated by, the cytoplasmic presence of REEs. In this context, it is interesting to note that in a recent proteomic approach, we found that besides PedE and PedH, additional proteins of unknown function show differential abundance in response to La 3+ availability (Wehrmann et al, 2019). It will hence be interesting to find out, whether any of these proteins is required for PedH function.…”

“…We have recently reported that P. putida KT2440 is capable of using rare earth elements (REEs) of the lanthanide series (Ln 3+ ) when growing on several alcoholic substrates (Wehrmann et al, 2017, 2019). Under these conditions, the cells use the pyrroloquinoline quinone (PQQ)-dependent ethanol dehydrogenase (EDH) PedH, to catalyze their initial oxidation within the periplasm.…”

Rare earth element (REE)-dependent growth of Pseudomonas putida KT2440 depends on the ABC-transporter PedA1A2BC and is influenced by iron availability

“…It is however possible that additional genes/proteins required for PedH-dependent growth rely on, or are regulated by, the cytoplasmic presence of REEs. In this context it is interesting to note that in a recent proteomic approach, we found that besides PedE and PedH, additional proteins of unknown function show differential abundance in response to La 3+ availability (Wehrmann et al, 2019). It will hence be interesting to find out in future studies whether any of these proteins is required for PedH function.…”

Insights into an alternative pathway for glycerol metabolism in a glycerol kinase deficientPseudomonas putidaKT2440

“…Analysis of the KT2440 genome revealed 5 further dehydrogenases that could be responsible for glycerate production in the mutant strain, these enzymes were between 2 and 2.7 fold upregulated in the transcriptomic analysis. A companion study to this work, which investigated the physiological responses of P. putida KT2440 towards rare earth elements during growth with different growth substrates, identified that two periplasmic PQQ-dependent alcohol dehydrogenases that are essential to initiate an alternative glycerol pathway via the oxidation of glycerol to glyceraldehyde (34). No significant difference in the expression of these PQQ-dependent dehydrogenases was observed in the transcriptomic analysis.…”