title: Overexpression and characterization of lycopene cyclase (CrtY) from marine bacterium Paracoccus haeundaensis.
abstract: Lycopene cyclase converts lycopene to beta-carotene by catalyzing the formation of two beta-rings at each end of the linear carotene structure. This reaction takes place as a two-step reaction in which both sides of of the lycopene molecule are cyclized into beta-carotene rings via the monocyclic gamma-carotene as an intermediate. The crtY gene coding for lycopene cyclase from Paracoccus haeundaensis consists of 1,158 base pairs encoding 386 amino acids residues. An expression plasmid containing the crtY gene (pET44a-CrtY) was constructed and expressed in Escherichia coli, and produced a recombinant protein of approximately 43 kDa, corresponding to the molecular mass of lycopene cyclase. The expressed protein was purified to homogeneity by His-tag affinity chromatography and showed enzymatic activity corresponding to lycopene cyclase. We also determined the lycopene substrate specificity and NADPH cofactor requirements of the purified protein. The Km values for lycopene and NADPH were 3.5 microM and 2 mM, respectively. The results obtained from this study will provide a wider base of knowledge on the enzyme characterization of lycopene cyclase at the molecular level.
authors: Department of Microbiology, Pukyong National University, Busan, Korea.
authors: Tae Hyug
authors: TH
authors: Jeong
authors: Keunho
authors: K
authors: Ji
authors: Young Tae
authors: YT
authors: Kim
mesh_terms: D002338:Carotenoids / Q000378:metabolism; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D003067:Coenzymes / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D019753:Intramolecular Lyases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D000077276:Lycopene; D008970:Molecular Weight; D009249:NADP / Q000378:metabolism; D010230:Paracoccus / Q000201:enzymology* / Q000235:genetics; D010957:Plasmids; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D013379:Substrate Specificity; D019207:beta Carotene / Q000378:metabolism
chemical_list: D003067:Coenzymes; D011994:Recombinant Proteins; D019207:beta Carotene; D002338:Carotenoids; D009249:NADP; D019753:Intramolecular Lyases; C070925:lycopene cyclase-isomerase; D000077276:Lycopene