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Lanthanides (Ln) are a new group of life metals, and many questions remain regarding how they are acquired and used in biology. Methylotrophic bacteria can acquire, transport, biomineralize, and use Ln as part of a cofactor complex with pyrroloquinoline quinone (PQQ) in alcohol dehydrogenases. For most methylotrophic bacteria use is restricted to the light Ln, which range from lanthanum to samarium (atomic numbers 57–62). Understanding how the cell differentiates between light and heavy Ln, and the impacts of these metals on the metabolic network, will advance the field of Ln biochemistry and give insights into enzyme catalysis, stress homeostasis, and metal biomineralization and compartmentalization. We report robust methanol growth with the heavy Ln gadolinium by a genetic variant of the model methylotrophic bacterium Methylorubrum extorquens AM1, named evo -HLn, for “ evo lved for H eavy L antha n ides.” A non-synonymous single nucleotide polymorphism in a cytosolic hybrid histidine kinase/response regulator allowed for sweeping transcriptional alterations to heavy metal stress response, methanol oxidation, and central metabolism. Increased expression of genes for Ln acquisition and uptake, production of the Ln-chelating lanthanophore, PQQ biosynthesis, and phosphate transport and metabolism resulted in gadolinium hyperaccumulation of 36-fold with a trade-off for light Ln accumulation. Gadolinium was hyperaccumulated in an enlarged acidocalcisome-like compartment. This is the first evidence of a bacterial intracellular Ln-containing compartment that we name the “lanthasome.” Carotenoid and toblerol biosynthesis were also upregulated. Due to its unique capabilities, evo -HLn can be used to further magnetic resonance imaging (MRI) and bioremediation technologies. In this regard, we show that gadolinium hyperaccumulation was sufficient to produce MRI contrast in whole cells, and that evo -HLn was able to readily acquire the metal from the MRI contrast agent gadopentetic acid. Finally, hyperaccumulation of gadolinium, differential uptake of light and heavy Ln, increased PQQ levels, and phosphate transport provide new insights into strategies for Ln recovery.
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Lanthanide-dependent alcohol dehydrogenases require an essential aspartate residue for metal coordination and enzymatic function
The lanthanide elements (Ln3), those with atomic numbers57–63 (excluding promethium, Pm3), form a cofactor complexwith pyrroloquinoline quinone (PQQ) in bacterial XoxF meth-anol dehydrogenases (MDHs) and ExaF ethanol dehydroge-nases (EDHs), expanding the range of biological elements andopening novel areas of metabolism and ecology. Other MDHs,known as MxaFIs, are related in sequence and structure to theseproteins, yet they instead possess a Ca2-PQQ cofactor. Animportant missing piece of the Ln3puzzle is defining what fea-tures distinguish enzymes that use Ln3-PQQ cofactors fromthose that do not. Here, using XoxF1 MDH from the modelmethylotrophic bacteriumMethylorubrum extorquensAM1, weinvestigated the functional importance of a proposed lantha-nide-coordinating aspartate residue. We report two crystalstructures of XoxF1, one with and another without PQQ, bothwith La3bound in the active-site region and coordinated byAsp320. Using constructs to produce either recombinant XoxF1or its D320A variant, we show that Asp320is needed forin vivocatalytic function,in vitroactivity, and La3coordination.XoxF1 and XoxF1 D320A, when produced in the absence ofLa3, coordinated Ca2but exhibited little or no catalytic activ-ity. We also generated the parallel substitution in ExaF to pro-duce ExaF D319S and found that this variant loses the capacityfor efficient ethanol oxidation with La3. These results provideevidence that a Ln3-coordinating aspartate is essential for theenzymatic functions of XoxF MDHs and ExaF EDHs, supportingthe notion that sequences of these enzymes, and the genes thatencode them, are markers for Ln3metabolism.
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- Award ID(s):
- 1807073
- PAR ID:
- 10233783
- Date Published:
- Journal Name:
- The journal of biological chemistry
- Volume:
- 295
- Issue:
- 24
- ISSN:
- 1067-8816
- Page Range / eLocation ID:
- 8272-8284
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.1074/jbc.RA120.013227
