Engineering nonribosomal peptide synthetases (NRPSs) has been a “holy grail” in synthetic biology due to their modular nature and limited understanding of catalytic mechanisms. Here, we reported a computational redesign of the “gate-keeper” adenylation domain of the model NRPS-like enzyme carboxylic acid reductases (CARs) by using approximate mechanism-based geometric criteria and the Rosetta energy score. Notably, MabCAR3 mutants ACA-1 and ACA-4 displayed a remarkable improvement in catalytic efficiency (kcat/KM) for 6-aminocaproic acid, up to 101-fold. Furthermore, G418K exhibited an 86-fold enhancement in substrate specificity for adipic acid compared to 6-aminocaproic acid. Our work provides not only promising biocatalysts for nylon monomer biosynthesis but also a strategy for efficient NRPSs engineering.@en

Handling excess sludge produced by wastewater treatment is a common problem worldwide. Due to limited space available in landfills, as well as difficulties involved in using excess sludge in agriculture, there is a need for alternative disposal methods. Although anaerobic digestion (AD) is widely used in processing sludge, only partial energy recovery from methane and sludge volume reduction can be achieved, resulting in a substantial amount of sludge remaining, which needs to be disposed of. Direct incineration after sludge drying is one possible option, a practice that is already in place in some cities in China. A comparison between direct incineration and conventional AD (with or without pretreatment by thermal hydrolysis) has to be made with respect to the energy balance and investment & operational (I & O) costs. This comparison reveals direct incineration to have the lowest energy deficit and I & O costs. Therefore, it is expected that direct incineration without AD will become the preferred sustainable approach to handling sludge.

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