title: Bacopa monniera recombinant mevalonate diphosphate decarboxylase: Biochemical characterization.
abstract: Mevalonate diphosphate decarboxylase (MDD; EC 4.1.1.33) is an important enzyme in the mevalonic acid pathway catalyzing the Mg(2+)-ATP dependant decarboxylation of mevalonate 5-diphosphate (MVAPP) to isopentenyl diphosphate (IPP). Bacopa monniera recombinant MDD (BmMDD) protein was overexpressed in Escherichia coli BL21 (DE3) strain and purified to apparent homogeneity. Km and Vmax for MVAPP were 144 μM and 52 U mg(-1) respectively. The values of turnover (kcat) and kcat/Km for mevalonate 5-diphosphate were determined to be 40s(-1) and 2.77×10(5) M(-1) s(-1) and kcat and kcat/Km values for ATP were found to be 30 s(-1) and 2.20×10(4) M(-1) s(-1), respectively. pH activity profile indicated the involvement of carboxylate ion, lysine and arginine for the activity of enzyme. The apparent activation energy for the BmMDD catalyzed reaction was 12.7 kJ mol(-1). Optimum pH and temperature for the forward reaction was found to be 8.0 and 45 °C. The enzyme was most stable at pH 7 at 20 °C with the deactivation rate constant (Kd(*)) of 1.69×10(-4) and half life (t1/2) of 68 h. The cation studies suggested that BmMDD is a cation dependant enzyme and optimum activity was achieved in the presence of Mg(2+).
authors.affiliation: Plant Tissue Culture Division, CSIR-National Chemical Laboratory, Dr. Homi Bhabha Road, Pune, Maharashtra, India.
authors.forename: Shakeel J
authors.initials: SJ
authors.lastname: Abbassi
authors.affiliation: Plant Tissue Culture Division, CSIR-National Chemical Laboratory, Dr. Homi Bhabha Road, Pune, Maharashtra, India.
authors.forename: Rishi K
authors.initials: RK
authors.lastname: Vishwakarma
authors.affiliation: Plant Tissue Culture Division, CSIR-National Chemical Laboratory, Dr. Homi Bhabha Road, Pune, Maharashtra, India.
authors.forename: Parth
authors.initials: P
authors.lastname: Patel
authors.affiliation: Plant Tissue Culture Division, CSIR-National Chemical Laboratory, Dr. Homi Bhabha Road, Pune, Maharashtra, India.
authors.forename: Uma
authors.initials: U
authors.lastname: Kumari
authors.affiliation: Plant Tissue Culture Division, CSIR-National Chemical Laboratory, Dr. Homi Bhabha Road, Pune, Maharashtra, India. Electronic address: bm.khan@ncl.res.in.
authors.forename: Bashir M
authors.initials: BM
authors.lastname: Khan
mesh_terms: D034681:Bacopa / Q000737:chemistry* / Q000201:enzymology; D002262:Carboxy-Lyases; D002413:Cations, Divalent; D003001:Cloning, Molecular; D057075:Enzyme Assays; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D045782:Hemiterpenes / Q000737:chemistry* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008274:Magnesium / Q000737:chemistry*; D008798:Mevalonic Acid / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D008970:Molecular Weight; D009943:Organophosphorus Compounds / Q000737:chemistry* / Q000378:metabolism; D010940:Plant Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013696:Temperature
chemical_list: D002413:Cations, Divalent; D045782:Hemiterpenes; D009943:Organophosphorus Compounds; D010940:Plant Proteins; D011994:Recombinant Proteins; C004809:isopentenyl pyrophosphate; C046285:5-diphosphomevalonic acid; D002262:Carboxy-Lyases; C022503:pyrophosphomevalonate decarboxylase; D008274:Magnesium; D008798:Mevalonic Acid
keywords: Mevalonate diphosphate decarboxylase; Phylogenetic analysis; Stability; pH activity profile