title: A novel mutation, D404N, in the connection subdomain of reverse transcriptase of HIV-1 CRF08_BC subtype confers cross-resistance to NNRTIs.
abstract: OBJECTIVE
  Growing evidence suggests that mutations in the connection domain of the HIV-1 reverse transcriptase (RT) can contribute to viral resistance to RT inhibitors. This work was designed to determine the effects of a novel mutation, D404N, in the connection subdomain of RT of HIV-1 CRF08_BC subtype on drug resistance, viral replication capacity (RC) and RT activity.
  
  
  METHODS
  Mutation D404N, alone or together with the other reported mutations, was introduced into an HIV-1 CRF08_BC subtype infectious clone by site-directed mutagenesis. Viral susceptibility to nine RT inhibitors, viral RC and the DNA polymerase activity of viral RT of the constructed virus mutants were investigated. A modelling study using the server SWISS-MODEL was conducted to explore the possible structure-related drug resistance mechanism of the mutation D404N.
  
  
  RESULTS
  Single mutations D404N and H221Y conferred low-level resistance to nevirapine, efavirenz, rilpivirine and zidovudine. Double mutations Y181C/D404N and Y181C/H221Y significantly reduced susceptibility to NNRTIs. The most pronounced resistance to NNRTIs was observed with the triple mutation Y181C/D404N/H221Y. Virus containing D404N as the only mutation displayed ∼50% RC compared with the WT virus. The modelling study suggested that the D404N mutation might abolish the hydrogen bonds between residues 404 and K30 in p51 or K431 in p66, leading to impaired RT subunit structure and enhanced drug resistance.
  
  
  CONCLUSIONS
  These results indicate that D404N is a novel NNRTI-associated mutation in the HIV-1 subtype CRF08_BC and provides information valuable for the monitoring of clinical RTI resistance.
authors: Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong, SAR, China.
authors: Xiao-Min
authors: XM
authors: Zhang
authors: Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong, SAR, China.
authors: Hao
authors: H
authors: Wu
authors: Biosafety Level-3 Laboratory, School of Public Health and Tropical Medicine, Southern Medical University, Guangzhou, China.
authors: Qiwei
authors: Q
authors: Zhang
authors: Department of Biomedical Sciences, City University of Hong Kong, Hong Kong, SAR, China.
authors: Terrence Chi-Kong
authors: TC
authors: Lau
authors: Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong, SAR, China.
authors: Hin
authors: H
authors: Chu
authors: Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong, SAR, China.
authors: Zhi-Wei
authors: ZW
authors: Chen
authors: Department of Biochemistry, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong, SAR, China.
authors: Dong-Yan
authors: DY
authors: Jin
authors: Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong, SAR, China bzheng@hkucc.hku.hk.
authors: Bo-Jian
authors: BJ
authors: Zheng
mesh_terms: D024882:Drug Resistance, Viral; D054303:HIV Reverse Transcriptase / Q000235:genetics* / Q000378:metabolism*; D015497:HIV-1 / Q000187:drug effects* / Q000201:enzymology*; D006801:Humans; D008826:Microbial Sensitivity Tests; D008958:Models, Molecular; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D020125:Mutation, Missense; D011487:Protein Conformation; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology*; D048348:Reverse Transcription
chemical_list: D050505:Mutant Proteins; D018894:Reverse Transcriptase Inhibitors; C514824:reverse transcriptase, Human immunodeficiency virus 1; D054303:HIV Reverse Transcriptase
keywords: drug susceptibility; modelling; replication capacity
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