Nanobodies and chemical cross-links advance the structural and functional analysis of PI3Kα.
Jonathan R HartXiao LiuChen PanAnyi LiangLynn UenoYingna XuAlexandra QuezadaXinyu ZouSu YangQingtong ZhouSteve SchoonoogheGholamreza Hassanzadeh-GhassabehTian XiaWenqing ShuiDehua YangPeter K VogtMing-Wei WangPublished in: Proceedings of the National Academy of Sciences of the United States of America (2022)
Nanobodies and chemical cross-linking were used to gain information on the identity and positions of flexible domains of PI3Kα. The application of chemical cross-linking mass spectrometry (CXMS) facilitated the identification of the p85 domains BH, cSH2, and SH3 as well as their docking positions on the PI3Kα catalytic core. Binding of individual nanobodies to PI3Kα induced activation or inhibition of enzyme activity and caused conformational changes that could be correlated with enzyme function. Binding of nanobody Nb3-126 to the BH domain of p85α substantially improved resolution for parts of the PI3Kα complex, and binding of nanobody Nb3-159 induced a conformation of PI3Kα that is distinct from known PI3Kα structures. The analysis of CXMS data also provided mechanistic insights into the molecular underpinning of the flexibility of PI3Kα.
Keyphrases
- molecular dynamics simulations
- mass spectrometry
- high glucose
- diabetic rats
- molecular dynamics
- single molecule
- dna binding
- high resolution
- drug induced
- oxidative stress
- healthcare
- liquid chromatography
- endothelial cells
- electronic health record
- big data
- machine learning
- crystal structure
- transcription factor
- health information
- gas chromatography
- deep learning