Since the discovery of the biological relevance of rare earth elements (REEs) for numerous different bacteria, questions concerning the advantages of REEs in the active sites of methanol dehydrogenases (MDHs) over calcium(II) and of why bacteria prefer light REEs have been a subject of debate. Here we report the cultivation and purification of the strictly REE‐dependent methanotrophic bacterium Methylacidiphilum fumariolicum SolV with europium(III), as well as structural and kinetic analyses of the first methanol dehydrogenase incorporating Eu in the active site. Crystal structure determination of the Eu‐MDH demonstrated that overall no major structural changes were induced by conversion to this REE. Circular dichroism (CD) measurements were used to determine optimal conditions for kinetic assays, whereas inductively coupled plasma mass spectrometry (ICP‐MS) showed 70 % incorporation of Eu in the enzyme. Our studies explain why bacterial growth of SolV in the presence of Eu3+ is significantly slower than in the presence of La3+/Ce3+/Pr3+: Eu‐MDH possesses a decreased catalytic efficiency. Although REEs have similar properties, the differences in ionic radii and coordination numbers across the series significantly impact MDH efficiency.

Methanol dehydrogenases (MDH) have recently taken the spotlight with the discovery that a large portion of these enzymes in nature utilize lanthanides in their active sites. The kinetic parameters of these enzymes are determined with a spectrophotometric assay first described by Anthony and Zatman 55 years ago. This artificial assay uses alkylated phenazines, such as phenazine ethosulfate (PES) or phenazine methosulfate (PMS), as primary electron acceptors (EAs) and the electron transfer is further coupled to a dye. However, many groups have reported problems concerning the bleaching of the assay mixture in the absence of MDH and the reproducibility of those assays. Hence, the comparison of kinetic data among MDH enzymes of different species is often cumbersome. Using mass spectrometry, UV-Vis and electron paramagnetic resonance (EPR) spectroscopy, we show that the side reactions of the assay mixture are mainly due to the degradation of assay components. Light-induced demethylation (yielding formaldehyde and phenazine in the case of PMS) or oxidation of PES or PMS as well as a reaction with assay components (ammonia, cyanide) can occur. We suggest here a protocol to avoid these side reactions. Further, we describe a modified synthesis protocol for obtaining the alternative electron acceptor, Wurster's blue (WB), which serves both as EA and dye. The investigation of two lanthanide-dependent methanol dehydrogenases from Methylorubrum extorquens AM1 and Methylacidiphilum fumariolicum SolV with WB, along with handling recommendations, is presented.

Developmental processes, such as angiogenesis, are associated with a constant remodeling of the actin cytoskeleton in response to different mechanical stimuli. The mechanosensitive transcription factors MRTF-A (MKL1) and YAP (also known as YAP1) are important mediators of this challenging adaptation process. However, it is as yet unknown whether both pathways respond in an identical or in a divergent manner to a given microenvironmental guidance cue. Here, we use a micropatterning approach to dissect single aspects of cellular behavior in a spatiotemporally controllable setting. Using the exemplary process of angiogenesis, we show that cell-cell contacts and adhesive surface area are shared regulatory parameters of MRTF and YAP on rigid 2D surfaces. By analyzing MRTF and YAP under laminar flow conditions and during cell migration on dumbbell-shaped microstructures, we demonstrate that they exhibit different translocation kinetics. In conclusion, our work promotes the application of micropatterning techniques as a cell biological tool to study mechanosensitive signaling in the context of angiogenesis.

Effects of pH and temperature on the distribution of different pyrroloquinoline quinone (PQQ) species was investigated along with the spectroscopic properties of an europium(iii) and PQQ-containing methanol dehydrogenase.

Zusammenfassung Die Selten‐Erd‐Elemente haben Naturwissenschaftler schon seit Jahrzehnten fasziniert und ihnen Kopfzerbrechen bereitet. Diese Elemente sind heute aus unserem täglichen Leben nicht mehr wegzudenken. Das verdanken sie ihren einzigartigen physikochemischen Eigenschaften. Aktuelle Forschungsergebnisse zeigen zudem, dass manche dieser Elemente essentiell für methanotrophe Bakterien sind. Wir stehen erst am Anfang einer Entdeckungsreise in die faszinierende bioanorganische Chemie dieser Elemente und warten gespannt, welche Überraschungen sie noch für uns bereithalten.