Rahman Md Ajijur, Kaiser Frank, Jamshidi Shirin, Freitas Monteiro Marta, Rahman Khondaker Miraz, Mullany Peter, Roberts Adam P
Department of Microbial Diseases, University College London, 256 Gray's Inn Road, London, WC1X 8LD, UK.
Department of Pharmacy, University of Rajshahi, Rajshahi, 6205, Bangladesh.
Sci Rep. 2020 Nov 26;10(1):20709. doi: 10.1038/s41598-020-77377-4.
Antibiotic resistance poses an increasing threat to global health. To tackle this problem, the identification of principal reservoirs of antibiotic resistance genes (ARGs) plus an understanding of drivers for their evolutionary selection are important. During a PCR-based screen of ARGs associated with integrons in saliva-derived metagenomic DNA of healthy human volunteers, two novel variants of genes encoding a D-alanine-D-alanine ligase (ddl6 and ddl7) located within gene cassettes in the first position of a reverse integron were identified. Treponema denticola was identified as the likely host of the ddl cassettes. Both ddl6 and ddl7 conferred high level resistance to D-cycloserine when expressed in Escherichia coli with ddl7 conferring four-fold higher resistance to D-cycloserine compared to ddl6. A SNP was found to be responsible for this difference in resistance phenotype between the two ddl variants. Molecular dynamics simulations were used to explain the mechanism of this phenotypic change at the atomic scale. A hypothesis for the evolutionary selection of ddl containing integron gene cassettes is proposed, based on molecular docking of plant metabolites within the ATP and D-cycloserine binding pockets of Ddl.
抗生素耐药性对全球健康构成日益严重的威胁。为解决这一问题，识别抗生素耐药基因（ARGs）的主要储存库并了解其进化选择的驱动因素至关重要。在对健康人类志愿者唾液来源的宏基因组DNA中与整合子相关的ARGs进行基于PCR的筛选过程中，鉴定出了位于反向整合子首位基因盒内的两个编码D-丙氨酸-D-丙氨酸连接酶的新基因变体（ddl6和ddl7）。齿垢密螺旋体被确定为ddl基因盒的可能宿主。当在大肠杆菌中表达时，ddl6和ddl7均赋予对D-环丝氨酸的高水平抗性，与ddl6相比，ddl7赋予对D-环丝氨酸的抗性高四倍。发现一个单核苷酸多态性（SNP）导致了这两种ddl变体在抗性表型上的差异。分子动力学模拟用于在原子尺度上解释这种表型变化的机制。基于植物代谢物在Ddl的ATP和D-环丝氨酸结合口袋内的分子对接，提出了含ddl的整合子基因盒进化选择的假设。
