Chen, ZhenZhenChenRappert, SugimaSugimaRappertSun, JibinJibinSunZeng, An-PingAn-PingZeng2023-02-222023-02-222011-05-14Journal of Biotechnology 154 (4): 248-254 (2011)http://hdl.handle.net/11420/14870Deregulation of allosteric inhibition of enzymes is a challenge for strain engineering and has been achieved so far primarily by random mutation and trial-and-error. In this work, we used aspartokinase, an important allosteric enzyme for industrial amino acids production, to demonstrate a predictive approach that combines protein dynamics and evolution for a rational reengineering of enzyme allostery. Molecular dynamic simulation of aspartokinase III (AK3) from Escherichia coli and statistical coupling analysis of protein sequences of the aspartokinase family allowed to identify a cluster of residues which are correlated during protein motion and coupled during the evolution. This cluster of residues forms an interconnected network mediating the allosteric regulation, including most of the previously reported positions mutated in feedback insensitive AK3 mutants. Beyond these mutation positions, we have successfully constructed another twelve targeted mutations of AK3 desensitized toward lysine inhibition. Six threonine-insensitive mutants of aspartokinase I-homoserine dehydrogenase I (AK1-HD1) were also created based on the predictions. The proposed approach can be widely applied for the deregulation of other allosteric enzymes.en1873-486320114248254Elsevier ScienceAllosteric regulationAspartokinaseMolecular dynamics simulationStatistical coupling analysisBiowissenschaften, BiologieJournal Article10.1016/j.jbiotec.2011.05.00521609739Journal Article