Squalene-hopene cyclases (SHCs) are the biocatalytic pendant to asymmetric Brønsted-acid catalysis and thus comprise enormous synthetic potential. Nevertheless, their substrate scope is currently limited to terpenes. Herein, we present how to tailor the SHC's cation cage for an enantioselective semipinacol rearrangement of bicyclic allylic alcohols to produce valuable oxa-carbon spirocyclic compounds. Exploiting the subtle divergence of SHC active sites combined with structure-guided semirational engineering, we designed a biocatalyst with a high catalytic performance of ∼4500 TTN and excellent enantioselectivity of 99.5% enantiomeric excess (ee). In silico studies suggest that a broadened active site is pivotal for catalysis. This intricate cationic rearrangement is easily scalable, employing lyophilized cell powder in water. Furthermore, our substrate scope studies demonstrate the acceptance of diverse ring-sized substrates but also reveal that the naturally confined active site limits the function as a general "semipinacolase." This study showcases the ability to harness the SHC's cation cage to tap into the broader field of asymmetric Brønsted-acid catalysis.
Enzyme-based biocatalysis exhibits multiple advantages over inorganic catalysts, including the biocompatibility and the unchallenged specificity of enzymes towards their substrate. The recovery and repeated use of enzymes is essential for any realistic application in biotechnology, but is not easily achieved with current strategies. For this purpose, enzymes are often immobilized on inorganic scaffolds, which could entail a reduction of the enzymes’ activity. Here, we show that immobilization to a nano-scaled biological scaffold, a nanonetwork of end-to-end cross-linked M13 bacteriophages, ensures high enzymatic activity and at the same time allows for the simple recovery of the enzymes. The bacteriophages have been genetically engineered to express AviTags at their ends, which permit biotinylation and their specific end-to-end self-assembly while allowing space on the major coat protein for enzyme coupling. We demonstrate that the phages form nanonetwork structures and that these so-called nanonets remain highly active even after re-using the nanonets multiple times in a flow-through reactor.
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