Imaging Flow Cytometry for High-Throughput Phenotyping of Synthetic Cells

The reconstitution of basic cellular functions in micrometer-sized liposomes has led to a surge of interest in the construction of synthetic cells. Microscopy and flow cytometry are powerful tools for characterizing biological processes in liposomes with fluorescence readouts. However, applying each method separately leads to a compromise...

Clonal Amplification-Enhanced Gene Expression in Synthetic Vesicles

In cell-free gene expression, low input DNA concentration severely limits the phenotypic output, which may impair in vitro protein evolution efforts. We address this challenge by developing CADGE, a strategy that is based on clonal isothermal amplification of a linear gene-encoding dsDNA template by the minimal Φ29 replication machinery and...

Directed evolution of unspecific peroxygenase in organic solvents

Fungal unspecific peroxygenases (UPOs) are efficient biocatalysts that insert oxygen atoms into nonactivated C–H bonds with high selectivity. Many oxyfunctionalization reactions catalyzed by UPOs are favored in organic solvents, a milieu in which their enzymatic activity is drastically reduced. Using as departure point the UPO secretion...

A Pioneering Career in Catalysis: Manfred T. Reetz

In this invited Account, we highlight the enormous scientific breadth of our mentor Professor Manfred T. Reetz. It stretches from the development of organometallic reagents and transition metal catalysts to the adventurous idea of directed evolution of chemo-, stereo-, and regioselective enzymes, which he considered to be most important. We...

Roadmap to Building a Cell: An Evolutionary Approach

Laboratory synthesis of an elementary biological cell from isolated components may aid in understanding of the fundamental principles of life and will provide a platform for a range of bioengineering and medical applications. In essence, building a cell consists in the integration of cellular modules into system’s level functionalities...

Selective Synthesis of the Human Drug Metabolite 5′-Hydroxypropranolol by an Evolved Self-Sufficient Peroxygenase

Propranolol is a widely used beta-blocker that is metabolized by human liver P450 monooxygenases into equipotent hydroxylated human drug metabolites (HDMs). It is paramount for the pharmaceutical industry to evaluate the toxicity and activity of these metabolites, but unfortunately, their synthesis has hitherto involved the use of severe...