Pantothenate kinase-associated neurodegeneration (PKAN), a progressive neurodegenerative disorder, is associated with impairment of pantothenate kinase function. Pantothenate kinase is the first enzyme required for de novo synthesis of CoA, an essential metabolic cofactor. The pathophysiology of PKAN is not understood, and there is no cure to halt or reverse the symptoms of this devastating disease. Recently, we and others presented a PKAN Drosophila model, and we demonstrated that impaired function of pantothenate kinase induces a neurodegenerative phenotype and a reduced lifespan. We have explored this Drosophila model further and have demonstrated that impairment of pantothenate kinase is associated with decreased levels of CoA, mitochondrial dysfunction, and increased protein oxidation. Furthermore, we searched for compounds that can rescue pertinent phenotypes of the Drosophila PKAN model and identified pantethine. Pantethine feeding restores CoA levels, improves mitochondrial function, rescues brain degeneration, enhances locomotor abilities, and increases lifespan. We show evidence for the presence of a de novo CoA biosynthesis pathway in which pantethine is used as a precursor compound. Importantly, this pathway is effective in the presence of disrupted pantothenate kinase function. Our data suggest that pantethine may serve as a starting point to develop a possible treatment for PKAN.coenzyme A | mitochondria | PKAN | oxidative stress | lifespan

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The use of enzymes to interfere with quorum sensing represents an attractive strategy to fight bacterial infections. We used PvdQ, an effective quorum-quenching enzyme from Pseudomonas aeruginosa, as a template to generate an acylase able to effectively hydrolyze C8-HSL, the major communication molecule produced by the Burkholderia species. We discovered that the combination of two single mutations leading to variant PvdQ Lα146W,Fβ24Y conferred high activity toward C8-HSL. Exogenous addition of PvdQ Lα146W,Fβ24Y dramatically decreased the amount of C8-HSL present in Burkholderia cenocepacia cultures and inhibited a quorum sensing-associated phenotype. The efficacy of this PvdQ variant to combat infections in vivo was further confirmed by its ability to rescue Galleria mellonella larvae upon infection, demonstrating its potential as an effective agent toward Burkholderia infections. Kinetic analysis of the enzymatic activities toward 3-oxo-C12-L-HSL and C8-L-HSL corroborated a substrate switch. This work demonstrates the effectiveness of quorumquenching acylases as potential novel antimicrobial drugs. In addition, we demonstrate that their substrate range can be easily switched, thereby paving the way to selectively target only specific bacterial species inside a complex microbial community.computational design | enzyme engineering | antibiotic | cystic fibrosis

eThe iron binding siderophore pyoverdine constitutes a major adaptive factor contributing to both virulence and survival in fluorescent pseudomonads. For decades, pyoverdine production has allowed the identification and classification of fluorescent and nonfluorescent pseudomonads. Here, we demonstrate that PvdP, a periplasmic enzyme of previously unknown function, is a tyrosinase required for the maturation of the pyoverdine chromophore in Pseudomonas aeruginosa. PvdP converts the nonfluorescent ferribactin, containing two iron binding groups, into a fluorescent pyoverdine, forming a strong hexadentate complex with ferrous iron, by three consecutive oxidation steps. PvdP represents the first characterized member of a small family of tyrosinases present in fluorescent pseudomonads that are required for siderophore maturation and are capable of acting on large peptidic substrates.

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Terpenoids belong to the largest class of natural compounds and are produced in all living organisms. The isoprenoid skeleton is based on assembling of C5 building blocks, but the biosynthesis of a great variety of terpenoids ranging from monoterpenoids to polyterpenoids is not fully understood today. Terpenoids play a fundamental role in human nutrition, cosmetics, and medicine. In the past 10 years, many metabolic engineering efforts have been undertaken in plants but also in microorganisms to improve the production of various terpenoids like artemisinin and paclitaxel. Recently, inverse metabolic engineering and combinatorial biosynthesis as main strategies in synthetic biology have been applied to produce high-cost natural products like artemisinin and paclitaxel in heterologous microorganisms. This review describes the recent progresses made in metabolic engineering of the terpenoid pathway with particular focus on fundamental aspects of host selection, vector design, and system biotechnology.

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Pyoverdine biosynthesis in fluorescent Pseudomonas spp. and especially in the opportunistic human pathogen Pseudomonas aeruginosa has been extensively studied. The acylase PvdQ is required for a maturation step in pyoverdine biosynthesis but also has been proven to be effective in degrading long-chain N-acyl homoserine lactones (AHLs). These molecules are used as quorum-sensing molecules by Gram-negative bacteria such as Pseudomonads themselves. Interestingly, the pvdQ gene is part of a pyoverdine cluster in P. aeruginosa and P. syringae but not in other fluorescent Pseudomonas spp. In this study we have compared the activities of PvdQ orthologues from various species and provide evidence for conserved functions in Pseudomonas fluorescens PfO-1, P. putida KT2440 and P. aeruginosa PA14. Despite large differences in genomic organization, expression of each of these pvdQ orthologues is regulated by iron availability. Moreover, PvdQ and its orthologues have conserved substrate specificity for AHLs and play a role in pyoverdine production in all tested Pseudomonas species. These data strongly suggest that the role of PvdQ in pyoverdine biosynthesis is conserved among Pseudomonas spp., while the control that PvdQ exerts in P. aeruginosa over its own quorum-sensing signals seems to be unique to this bacterium.