title: Identification and characterization of a pantothenate kinase (PanK-sp) from streptomyces peucetius ATCC 27952.
abstract: Pantothenate kinase (PanK) catalyzes the first step in the biosynthesis of the essential and ubiquitous cofactor coenzyme A (CoA) in all organisms. Here, we report the identification, cloning, and characterization of panK-sp from Streptomyces peucetius ATCC 27952. The gene encoded a protein of 332 amino acids with a calculated molecular mass of 36.8 kDa and high homology with PanK from S. avermitilis and S. coelicolor A3(2). To elucidate the putative function of PanK-sp, it was cloned into pET32a(+) to construct pPKSP32, and the PanK-sp was then expressed in E. coli BL21(DE3) as a His-tag fusion protein and purified by immobilized metal affinity chromatography. The enzyme assay of PanK-sp was carried out as a coupling assay. The gradual decrease in NADH concentration with time clearly indicated the phosphorylating activity of PanK-sp. Furthermore, the ca. 1.4-fold increase of DXR and the ca. 1.5-fold increase of actinorhodin by in vivo overexpression of panK-sp, constructed in pIBR25 under the control of a strong ermE* promoter, established its positive role in secondary metabolite production from S. peucetius and S. coelicolor, respectively.
authors.affiliation: Institute of Biomolecule Reconstruction (iBR), Department of Pharmaceutical Engineering, Sun Moon University, Asansi 336-708, Korea.
authors.forename: Ariungerel
authors.initials: A
authors.lastname: Mandakh
authors.forename: Narayan Prasad
authors.initials: NP
authors.lastname: Niraula
authors.forename: Eung Pil
authors.initials: EP
authors.lastname: Kim
authors.forename: Jae Kyung
authors.initials: JK
authors.lastname: Sohng
mesh_terms: D000880:Anthraquinones / Q000378:metabolism; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D003067:Coenzymes / Q000378:metabolism; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009243:NAD / Q000378:metabolism; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013302:Streptomyces / Q000201:enzymology*; D000095622:Benzoisochromanequinones
chemical_list: D000880:Anthraquinones; D003067:Coenzymes; D004269:DNA, Bacterial; D009243:NAD; D017853:Phosphotransferases (Alcohol Group Acceptor); D011994:Recombinant Proteins; C023088:pantothenate kinase; C013390:actinorhodin; D000095622:Benzoisochromanequinones