Solvent concentration at 50% protein unfolding may reform enzyme stability ranking and process window identification.

As water miscible organic co-solvents are often required for enzyme reactions to improve e.g., the solubility of the substrate in the aqueous medium, an enzyme is required which displays high stability in the presence of this co-solvent. Consequently, it is of utmost importance to identify the most suitable enzyme or the appropriate reaction conditions. Until now, the melting temperature is used in general as a measure for stability of enzymes. The experiments here show, that the melting temperature does not correlate to the activity observed in the presence of the solvent. As an alternative parameter, the concentration of the co-solvent at the point of 50% protein unfolding at a specific temperature T in short c U 50 T is introduced. Analyzing a set of ene reductases, c U 50 T is shown to indicate the concentration of the co-solvent where also the activity of the enzyme drops fastest. Comparing possible rankings of enzymes according to melting temperature and c U 50 T reveals a clearly diverging outcome also depending on the specific solvent used. Additionally, plots of c U 50 versus temperature enable a fast identification of possible reaction windows to deduce tolerated solvent concentrations and temperature.

Redox-Active Triazole-Derived Mesoionic Imines with Ferrocenyl Substituents and their Metal Complexes: Directed Hydrogen-Bonding, Unusual C-H Activation and Ion-Pair Formation.

We present herein the synthesis, characterization and complexation of ferrocenyl-substituted MIIs (mesoionic imines) and their metal complexes. In the free MIIs, strong hydrogen bonding interactions are observed between the imine-N and the C-H bonds of the ferrocenyl substituents both in the solid state and in solution. The influence of this hydrogen bonding is so strong that complexation of the MIIs with [IrCp*Cl2]2 yields unique six-membered iridacycles via C-H-activation of the corresponding C-H-site at the Fc-substituent and not the Ph-substituent. This result is in contrast to previous reports in which always a preferential C-H activation at the phenyl substituent is observed in competitive reactions in the presence of ferrocenyl substituents. The corresponding Ir complexes formed after in-situ halide exchange reaction exist in either [Ir-I] contact or as [Ir]+I- solvent separated ion-pairs depending on the solvent polarity. The iodide coordinated and solvent separated ion-pairs display drastically different physical properties. The TEP (Tolman-electronic-parameter) of these ligands was determined and lines up with previously reported MII-ligands. The redox properties were investigated by a combination of electrochemical and spectroelectrochemical methods. We show here how non-covalent interactions can have a drastic influence on the physical and chemical properties of these new class of compounds.