title: Mixed Inhibition of cPEPCK by Genistein, Using an Extended Binding Site Located Adjacent to Its Catalytic Cleft.
abstract: Cytosolic phosphoenolpyruvate carboxykinase (cPEPCK) is a critical enzyme involved in gluconeogenesis, glyceroneogenesis and cataplerosis. cPEPCK converts oxaloacetic acid (OAA) into phosphoenol pyruvate (PEP) in the presence of GTP. cPEPCK is known to be associated with type 2 diabetes. Genistein is an isoflavone compound that shows anti-diabetic and anti-obesitic properties. Experimental studies have shown a decrease in the blood glucose level in the presence of genistein by lowering the functional activity of cPEPCK, an enzyme of gluconeogenesis. Using computational techniques such as molecular modeling, molecular docking, molecular dynamics simulation and binding free energy calculations, we identified cPEPCK as a direct target of genistein. We studied the molecular interactions of genistein with three possible conformations of cPEPCK-unbound cPEPCK (u_cPEPCK), GTP bound cPEPCK (GTP_cPEPCK) and GDP bound cPEPCK (GDP_cPEPCK). Binding of genistein was also compared with an already known cPEPCK inhibitor. We analyzed the interactions of genistein with cPEPCK enzyme and compared them with its natural substrate (OAA), product (PEP) and known inhibitor (3-MPA). Our results demonstrate that genistein uses the mechanism of mixed inhibition to block the functional activity of cPEPCK and thus can serve as a potential anti-diabetic and anti-obesity drug candidate. We also identified an extended binding site in the catalytic cleft of cPEPCK which is used by 3-MPA to inhibit cPEPCK non-competitively. We demonstrate that extended binding site of cPEPCK can further be exploited for designing new drugs against cPEPCK.
authors.affiliation: Department of Biochemical Engineering and Biotechnology, Indian Institute of Technology (IIT) Delhi, Hauz Khas, New Delhi, India.
authors.forename: Shashank Prakash
authors.initials: SP
authors.lastname: Katiyar
authors.affiliation: Department of Biochemical Engineering and Biotechnology, Indian Institute of Technology (IIT) Delhi, Hauz Khas, New Delhi, India.
authors.forename: Arpit
authors.initials: A
authors.lastname: Jain
authors.affiliation: Department of Biochemical Engineering and Biotechnology, Indian Institute of Technology (IIT) Delhi, Hauz Khas, New Delhi, India.
authors.forename: Jaspreet Kaur
authors.initials: JK
authors.lastname: Dhanjal
authors.affiliation: Department of Biochemical Engineering and Biotechnology, Indian Institute of Technology (IIT) Delhi, Hauz Khas, New Delhi, India.
authors.forename: Durai
authors.initials: D
authors.lastname: Sundar
mesh_terms: D001665:Binding Sites; D002384:Catalysis; D019833:Genistein / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D008958:Models, Molecular; D019764:Phosphoenolpyruvate Carboxykinase (ATP) / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding
chemical_list: D019833:Genistein; C000597251:PCK2 protein, human; D019764:Phosphoenolpyruvate Carboxykinase (ATP)