Molecular insights into the catalytic promiscuity of a bacterial diterpene synthase.

Diterpene synthase VenA is responsible for assembling venezuelaene A with a unique 5-5-6-7 tetracyclic skeleton from geranylgeranyl pyrophosphate. VenA also demonstrates substrate promiscuity by accepting geranyl pyrophosphate and farnesyl pyrophosphate as alternative substrates. Herein, we report the crystal structures of VenA in both apo form and holo form in complex with a trinuclear magnesium cluster and pyrophosphate group. Functional and structural investigations on the atypical 115DSFVSD120 motif of VenA, versus the canonical Asp-rich motif of DDXX(X)D/E, reveal that the absent second Asp of canonical motif is functionally replaced by Ser116 and Gln83, together with bioinformatics analysis identifying a hidden subclass of type I microbial terpene synthases. Further structural analysis, multiscale computational simulations, and structure-directed mutagenesis provide significant mechanistic insights into the substrate selectivity and catalytic promiscuity of VenA. Finally, VenA is semi-rationally engineered into a sesterterpene synthase to recognize the larger substrate geranylfarnesyl pyrophosphate.

Progress in research and application development of surface display technology using Bacillus subtilis spores.

Bacillus subtilis is a widely distributed aerobic Gram-positive species of bacteria. As a tool in the lab, it has the advantages of nonpathogenicity and limited likelihood of becoming drug resistant. It is a probiotic strain that can be directly used in humans and animals. It can be induced to produce spores under nutrient deficiency or other adverse conditions. B. subtilis spores have unique physical, chemical, and biochemical characteristics. Expression of heterologous antigens or proteins on the surface of B. subtilis spores has been successfully performed for over a decade. As an update and supplement to previously published research, this paper reviews the latest research on spore surface display technology using B. subtilis. We have mainly focused on the regulation of spore coat protein expression, display and application of exogenous proteins, and identification of developing research areas of spore surface display technology.

Biocatalytic production of 2,5-furandicarboxylic acid: recent advances and future perspectives.

2,5-Furandicarboxylic acid (FDCA) is attracting increasing attention because of its potential applications as a sustainable substitute to petroleum-derived terephthalic acid for the production of bio-based polymers, such as poly(ethylene 2,5-furandicarboxylate) (PEF). Many catalytic methods have been developed for the synthesis of FDCA, including chemocatalysis, biocatalysis, photocatalysis, and electrocatalysis. Biocatalysis is a promising approach with advantages that include mild reaction condition, lower cost, higher selectivity, and environment amity. However, the biocatalytic production of FDCA has hardly been reviewed. To fully understand the current research developments, this review comprehensively considers the research progress on toxic effects and biodegradation of furan aldehydes, and then summarizes the latest achievements concerning the synthesis of FDCA from 5-hydroxymethylfurfural and other chemicals, such as 2-furoic acid and 5-methoxymethylfurfural. Our primary focus is on biocatalytic methods, including enzymatic catalysis (in vitro) and whole-cell catalysis (in vivo). Furthermore, future research directions and general developmental trends for more efficient biocatalytic production of FDCA are also proposed.