Diagram of the mode of action of DNA glycosylases TxnU2/U4 and LldU1/U5 cleaving TXNA/LLD-DNA adducts. Photo by Hangzhou Institute for Advanced Study, UCAS
WWW.CHINANEWS.COM, Hangzhou, February 25 (Huang Lingyi) On February 25, the reporter learned from the Hangzhou Institute for Advanced Study, UCAS that the Institute of Chemistry and Materials Science and the research group of Tang Gongli from the Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, cooperated with the research group of Brandt F. Eichman of Vanderbilt University. Their research result "Base excision repair system targeting DNA adducts of trioxacarcin/LL-D49194 antibiotics for self-resistance" was recently published in Nucleic Acids Research, an international academic journal. This study provides new clues for the solution of the clinical problem of resistance to DNA alkylating antibiotics and the development of anti-tumor drugs.
With the widespread use of antibiotics, the ensuing problem of antibiotic resistance is now one of the greatest health threats that humanity will need to face in the coming decades. The analysis of the mechanism of resistance to this class of antibiotics in this study could reveal the origin of resistance to Trioxacarcins (TXN) and LL-D49194 (LLD).
Base excision repair (BER) is a common damage repair pathway used to eliminate alkylated DNA in cells. DNA glycosylases, key proteins that identify and remove damaged bases, are the first step of the BER pathway. In recent years, based on studies on the self-resistance mechanisms of DNA-alkylating natural products, AlkD/YtkR2 and AlkZ/YcaQ are two families of DNA glycosylases in the antibiotic-producing strains that can recognize and repair DNA damage formed by DNA alkylates azinomycin B and yatakemycin, DNA alkylating compounds.
TXN and LLD are polycyclic aromatic polyketide natural products with highly complex structures, which contain multiple glycosylation groups allowing embedment into the DNA backbones and a highly active polycyclic-spiro epoxy three-membered ring structure. The two can be embedded into the major and minor DNA grooves and alkylated with guanine on the DNA chains to form stable DNA damage, realizing anti-malarial, anti-bacterial and anti-tumor activities.
Tang Gongli's research group has long been committed to researching the biosynthesis and resistance mechanism of TXNs, and successively clarified TXNs' initial elements and subsequent modification steps. Recently, they conducted an in-depth analysis of unknown functional genes in the gene clusters, from which they found four encoded DNA glycosylases (TxnU2, TxnU4, LldU1 and LldU5) conducting the alkylation damage repair of TXN A and LLD. These enzymes will provide self-resistance for TXN A and LLD producing strains.
It is reported that the amino acid sequence-based analysis of a protein's structure (using AlphaFold) shows that TxnU, LldU, and DNA glycosylase AlkZ/YcaQ are homologous; the in-vitro enzyme activity experiments prove that TxnU and LldU are single-functional DNA glycosylases acting on alkylguanine adducts at the N7 position. However, compared with homologous proteins and other DNA glycosylases that act on alkylpurine adducts, TxnU and LldU differ from each other regarding the substrate specificity and enzyme catalytic mechanism. TxnU/LldU can process the specific elimination of TXN A/LLD-DNA damage and is inactive against substrates of AlkD/YtkR2 and AlkZ/YcaQ, less stable alkylguanine adducts. Similarly, the TXN A/LLD-DNA adducts cannot be excised by any other alkylpurine DNA glycosylase. The results of docking modeling and point mutation experiments show that TxnU4 and LldU1 have unique catalytically active sequence motifs, from which their unique substrate specificities and catalytic mechanisms are explained. (End)
Executive Editor | Yuan Jingjing
