Functional Role of Lanthanides in Enzymatic Activity and Transcriptional Regulation of Pyrroloquinoline Quinone-Dependent Alcohol Dehydrogenases in <i>Pseudomonas putida</i> KT2440

The oxidation of alcohols and aldehydes is crucial for detoxification and efficient catabolism of various volatile organic compounds (VOCs). Thus, many Gram-negative bacteria have evolved periplasmic oxidation systems based on pyrroloquinoline quinone-dependent alcohol dehydrogenases (PQQ-ADHs) that are often functionally redundant. Here we report the first description and characterization of a lanthanide-dependent PQQ-ADH (PedH) in a nonmethylotrophic bacterium based on the use of purified enzymes from the soil-dwelling model organism <i>Pseudomonas putida</i> KT2440. PedH (PP_2679) exhibits enzyme activity on a range of substrates similar to that of its Ca²⁺-dependent counterpart PedE (PP_2674), including linear and aromatic primary and secondary alcohols, as well as aldehydes, but only in the presence of lanthanide ions, including La³⁺, Ce³⁺, Pr³⁺, Sm³⁺, or Nd³⁺ Reporter assays revealed that PedH not only has a catalytic function but is also involved in the transcriptional regulation of <i>pedE</i> and <i>pedH</i>, most likely acting as a sensory module. Notably, the underlying regulatory network is responsive to as little as 1 to 10 nM lanthanum, a concentration assumed to be of ecological relevance. The present study further demonstrates that the PQQ-dependent oxidation system is crucial for efficient growth with a variety of volatile alcohols. From these results, we conclude that functional redundancy and inverse regulation of PedE and PedH represent an adaptive strategy of <i>P. putida</i> KT2440 to optimize growth with volatile alcohols in response to the availability of different lanthanides.<b>IMPORTANCE</b> Because of their low bioavailability, lanthanides have long been considered biologically inert. In recent years, however, the identification of lanthanides as a cofactor in methylotrophic bacteria has attracted tremendous interest among various biological fields. The present study reveals that one of the two PQQ-ADHs produced by the model organism <i>P. putida</i> KT2440 also utilizes lanthanides as a cofactor, thus expanding the scope of lanthanide-employing bacteria beyond the methylotrophs. Similar to the system described in methylotrophic bacteria, a complex regulatory network is involved in lanthanide-responsive switching between the two PQQ-ADHs encoded by <i>P. putida</i> KT2440. We further show that the functional production of at least one of the enzymes is crucial for efficient growth with several volatile alcohols. Overall, our study provides a novel understanding of the redundancy of PQQ-ADHs observed in many organisms and further highlights the importance of lanthanides for bacterial metabolism, particularly in soil environments.