title: The Mechanism of ATP-Dependent Allosteric Protection of Akt Kinase Phosphorylation.
abstract: Kinases use ATP to phosphorylate substrates; recent findings underscore the additional regulatory roles of ATP. Here, we propose a mechanism for allosteric regulation of Akt1 kinase phosphorylation by ATP. Our 4.7-μs molecular dynamics simulations of Akt1 and its mutants in the ATP/ADP bound/unbound states revealed that ATP occupancy of the ATP-binding site stabilizes the closed conformation, allosterically protecting pT308 by restraining phosphatase access and key interconnected residues on the ATP→pT308 allosteric pathway. Following ATP→ADP hydrolysis, pT308 is exposed and readily dephosphorylated. Site-directed mutagenesis validated these predictions and indicated that the mutations do not impair PDK1 and PP2A phosphatase recruitment. We further probed the function of residues around pT308 at the atomic level, and predicted and experimentally confirmed that Akt1(H194R/R273H) double mutant rescues pathology-related Akt1(R273H). Analysis of classical Akt homologs suggests that this mechanism can provide a general model of allosteric kinase regulation by ATP; as such, it offers a potential avenue for allosteric drug discovery.
authors.affiliation: Department of Pathophysiology, Key Laboratory of Cell Differentiation and Apoptosis of Chinese Ministry of Education, Shanghai JiaoTong University, School of Medicine (SJTU-SM), Shanghai 200025, China.
authors.forename: Shaoyong
authors.initials: S
authors.lastname: Lu
authors.affiliation: Department of Biochemistry and Molecular Cell Biology & Shanghai Key Laboratory of Tumor Microenvironment and Inflammation, Shanghai JiaoTong University, School of Medicine (SJTU-SM), Shanghai 200025, China.
authors.forename: Rong
authors.initials: R
authors.lastname: Deng
authors.affiliation: Department of Pathophysiology, Key Laboratory of Cell Differentiation and Apoptosis of Chinese Ministry of Education, Shanghai JiaoTong University, School of Medicine (SJTU-SM), Shanghai 200025, China.
authors.forename: Haiming
authors.initials: H
authors.lastname: Jiang
authors.affiliation: Department of Pathophysiology, Key Laboratory of Cell Differentiation and Apoptosis of Chinese Ministry of Education, Shanghai JiaoTong University, School of Medicine (SJTU-SM), Shanghai 200025, China.
authors.forename: Huili
authors.initials: H
authors.lastname: Song
authors.affiliation: Department of Pathophysiology, Key Laboratory of Cell Differentiation and Apoptosis of Chinese Ministry of Education, Shanghai JiaoTong University, School of Medicine (SJTU-SM), Shanghai 200025, China.
authors.forename: Shuai
authors.initials: S
authors.lastname: Li
authors.affiliation: Department of Pathophysiology, Key Laboratory of Cell Differentiation and Apoptosis of Chinese Ministry of Education, Shanghai JiaoTong University, School of Medicine (SJTU-SM), Shanghai 200025, China.
authors.forename: Qiancheng
authors.initials: Q
authors.lastname: Shen
authors.affiliation: Department of Pathophysiology, Key Laboratory of Cell Differentiation and Apoptosis of Chinese Ministry of Education, Shanghai JiaoTong University, School of Medicine (SJTU-SM), Shanghai 200025, China.
authors.forename: Wenkang
authors.initials: W
authors.lastname: Huang
authors.affiliation: Cancer and Inflammation Program, Leidos Biomedical Research, Inc., Frederick National Laboratory, NCI, Frederick, MD 21702, USA; Department of Human Genetics and Molecular Medicine, Sackler School of Medicine, Sackler Institute of Molecular Medicine, Tel Aviv University, Tel Aviv 69978, Israel.
authors.forename: Ruth
authors.initials: R
authors.lastname: Nussinov
authors.affiliation: Department of Biochemistry and Molecular Cell Biology & Shanghai Key Laboratory of Tumor Microenvironment and Inflammation, Shanghai JiaoTong University, School of Medicine (SJTU-SM), Shanghai 200025, China. Electronic address: jianxiu.yu@gmail.com.
authors.forename: Jianxiu
authors.initials: J
authors.lastname: Yu
authors.affiliation: Department of Pathophysiology, Key Laboratory of Cell Differentiation and Apoptosis of Chinese Ministry of Education, Shanghai JiaoTong University, School of Medicine (SJTU-SM), Shanghai 200025, China. Electronic address: jian.zhang@sjtu.edu.cn.
authors.forename: Jian
authors.initials: J
authors.lastname: Zhang
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D000494:Allosteric Regulation; D001665:Binding Sites; D006367:HeLa Cells; D006801:Humans; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D054648:Protein Phosphatase 2 / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D000081382:Pyruvate Dehydrogenase Acetyl-Transferring Kinase
chemical_list: C000631429:PDK1 protein, human; D000081382:Pyruvate Dehydrogenase Acetyl-Transferring Kinase; D000255:Adenosine Triphosphate; C494918:AKT1 protein, human; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D054648:Protein Phosphatase 2