机构：[1]College of Animal Science and Technology, Jilin Agricultural University, Changchun, Jilin, China.[2]College of Veterinary Medicine, Jilin University, Changchun, Jilin, China.[3]Zhejiang Provincial Key Laboratory for Cancer Molecular Cell Biology, Life Sciences Institute, Zhejiang University, Hangzhou, Zhejiang, China.[4]College of Animal Science and Technology, Jilin Agricultural Science and Technology University, Changchun, Jilin, China.[5]Infectious Diseases and Pathogen Biology Center, Clinical Laboratory Department, First Hospital of Jilin University, Changchun, Jilin, China.[6]College of Chinese Medicine Materials, Jilin Agricultural University, Changchun, Jilin, China.[7]Faculty of Life and Health Sciences, Shenzhen Institute of Advanced Technology, Chinese Academy of Sciences, Shenzhen, Guangdong, China.[8]Inter disciplinary Center for Brain Information, The Brain Cognition and Brain Disease Institute, Shenzhen Institute of Advanced Technology, Chinese Academy of Sciences, Shenzhen, Guangdong, China.[9]Shenzhen-Hong Kong Institute of Brain Science-Shenzhen Fundamental Research Institutions, Shenzhen, Guangdong, China.深圳市康宁医院深圳医学信息中心[10]Jiangsu Co-Innovation Center for the Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou University, Yangzhou, Jiangsu, China.
Hypervirulent Klebsiella pneumoniae with capsular polysaccharides (CPSs) causes severe nosocomial- and community-acquired infections. Phage-derived depolymerases can degrade CPSs from K. pneumoniae to attenuate bacterial virulence, but their antimicrobial mechanisms and clinical potential are not well understood. In the present study, Klebsiella phage GH-K3-derived depolymerase Depo32 (encoded by gene gp32) was identified to exhibit high efficiency in specifically degrading the CPSs of K2 serotype K. pneumoniae. The cryo-electron microscopy structure of trimeric Depo32 at a resolution up to 2.32 Å revealed potential catalytic centers in the cleft of each of the two adjacent subunits. K. pneumoniae subjected to Depo32 became more sensitive to phagocytosis by RAW264.7 cells and activated the cells by the mitogen-activated protein kinase signaling pathway. In addition, intranasal inoculation with Depo32 (a single dose of 200 µg, 20 µg daily for 3 days, or in combination with gentamicin) rescued all C57BL/6J mice infected with a lethal dose of K. pneumoniae K7 without interference from its neutralizing antibody. In summary, this work elaborates on the mechanism by which Depo32 targets the degradation of K2 serotype CPSs and its potential as an antivirulence agent. IMPORTANCE Depolymerases specific to more than 20 serotypes of Klebsiella spp. have been identified, but most studies only evaluated the single-dose treatment of depolymerases with relatively simple clinical evaluation indices and did not reveal the anti-infection mechanism of these depolymerases in depth. On the basis of determining the biological characteristics, the structure of Depo32 was analyzed by cryo-electron microscopy, and the potential active center was further identified. In addition, the effects of Depo32 on macrophage phagocytosis, signaling pathway activation, and serum killing were revealed, and the efficacy of the depolymerase (single treatment, multiple treatments, or in combination with gentamicin) against acute pneumonia caused by Klebsiella pneumoniae was evaluated. Moreover, the roles of the active sites of Depo32 were also elucidated in the in vitro and in vivo studies. Therefore, through structural biology, cell biology, and in vivo experiments, this study demonstrated the mechanism by which Depo32 targets K2 serotype K. pneumoniae infection.
基金：
This work was financiallysupported through grants from the National Natural Science Foundation of China (grant Nos. 32072824, 32222083, U19A2038, 81802056, 32161133022, and 32102677), the Fundamental Research Funds for the Central Universities, the Natural Science Foundation of Jilin Province (No. YDZJ202201ZYTS449), the Natural Science Foundation of Shandong Province (ZR2020MC182), the Science and Technology Project of the Jilin Provincial Education Department (JJKH20210364KJ), and the China Postdoctoral Science Foundation (2021M691224).
语种：
外文
PubmedID：
中科院(CAS)分区：
出版当年[2022]版：
大类|1 区生物学
小类|1 区微生物学
最新[2025]版：
大类|2 区生物学
小类|3 区微生物学
第一作者：
第一作者机构：[1]College of Animal Science and Technology, Jilin Agricultural University, Changchun, Jilin, China.[2]College of Veterinary Medicine, Jilin University, Changchun, Jilin, China.
共同第一作者：
通讯作者：
通讯机构：[2]College of Veterinary Medicine, Jilin University, Changchun, Jilin, China.[10]Jiangsu Co-Innovation Center for the Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou University, Yangzhou, Jiangsu, China.
推荐引用方式(GB/T 7714)：
Cai Ruopeng,Ren Zhuolu,Zhao Rihong,et al.Structural biology and functional features of phage-derived depolymerase Depo32 on Klebsiella pneumoniae with K2 serotype capsular polysaccharides[J].Microbiology spectrum.2023,e0530422.doi:10.1128/spectrum.05304-22.
APA：
Cai Ruopeng,Ren Zhuolu,Zhao Rihong,Lu Yan,Wang Xinwu...&Gu Jingmin.(2023).Structural biology and functional features of phage-derived depolymerase Depo32 on Klebsiella pneumoniae with K2 serotype capsular polysaccharides.Microbiology spectrum,,
MLA：
Cai Ruopeng,et al."Structural biology and functional features of phage-derived depolymerase Depo32 on Klebsiella pneumoniae with K2 serotype capsular polysaccharides".Microbiology spectrum .(2023):e0530422
生物学