Reconstruction of a microbial TNT deep degradation system and its mechanism for reshaping microecology

This study is the first to use synthetic biological omics technology to analyze the molecular mechanism underlying deep degradation of TNT, to construct an artificial transformation system to create engineered Escherichia coli bacteria, and to use Bacillus subtilis as an expression host to explore the mechanism driving the reshaping of the deep degradation platform on microecology. Nitroreductase family protein, 2-oxoacid:acceptor oxidoreductase, NADPH-cytochrome P450 reductase, monooxygenase, ring-cleaving dioxygenase, and RraA family protein significantly participated in the reduction-hydroxylation-ring opening cleavage of TNT, achieving deep transformation of TNT to produce pyruvic acid and other products that entered the cellular metabolic cycle. The key toxic metabolic pathways of TNT, 2,4-diamino-6-nitrotoluene, 2,4,6-triaminotoluene, and 2,4,6-trihydroxytoluene are pantothenate and CoA biosynthesis. The engineered bacteria that impart TNT deep degradation ability regulate and optimize lipid, sugar, and amino acid metabolism to withstand stress. Engineered B. subtilis bacteria occupy ecological niches after repairing TNT-contaminated soil and water bodies while simultaneously recruiting a variety of microorganisms to reshape and positively regulate microecology. Key drivers for reshaping and optimization of microecological functions include ABC transporters and C/N/P/S functional cycles, together with a significant concomitant upregulation of the metabolic cycle of basic carbohydrates, nucleotides, and amino acids in the microecology.