|Table of Contents|

Cloning and Bioinformatic Analysis and Prokaryotic Expression of LaSPS Gene From Lepidium apetalum(PDF)

《北方园艺》[ISSN:1001-0009/CN:23-1247/S]

Issue:
2016年06
Page:
92-99
Research Field:
Publishing date:

Info

Title:
Cloning and Bioinformatic Analysis and Prokaryotic Expression of LaSPS Gene From Lepidium apetalum
Author(s):
ZHAO Le1; 2; MA Ligang1; 2; HAN Duwan1; FENG Weisheng1; 2; ZHENG Xiaoke1; 2
(1.School of Pharmacy，Henan University of Traditional Chinese Medicine，Zhengzhou，Henan 450046；2.Collaborative Innovation Center for Respiratory Disease Diagnosis and Treatment & Chinese Medicine Development of Henan Province，Zhengzhou，Henan 450046)
Keywords:
Lepidium apetalum; solanesyl diphosphate synthase; gene cloning; sequence analysis; prokaryotic expression
PACS:
-
DOI:
10.11937/bfyy.201606023
Abstract:
Taking leaves of Lepidium apetalum as experimental material，using PCR method，the open reading frame (ORF) of LaSPS gene was amplified，and the prokaryotic expression vector pET-32a-LaSPS was constructed and then Escherichia coli BL21 (DE3) cells were transformed with the plasmid pET-32a-LaSPS in order to express recombinant LaSPS protein.The results indicated that the ORF of LaSPS gene was 1 269 bp (GenBank accession number KT318734)， which encoded a protein of 422 amino acid residues.The LaSPS protein which contained solanesyl diphosphate synthase domain was the member of ‘polyprenyl diphosphate synthase’ family.Bioinformatic analysis indicated that LaSPS protein which located in chloroplast had no transmembrane domain and signal peptide.The multiple alignment and phylogenetic analysis indicated that LaSPS protein showed the highest homology，91% similarity，with AtSPS2 protein from Arabidopsis thaliana.The prokaryotic expression vector pET-32a-LaSPS was constructed and the recombinant LaSPS protein was successfully expressed in E.coli BL21 (DE3) cells.This study provided the foundation for follow-up research of its function in plastoquinoe biosynthesis pathway.

References:

 

[1]JONES M O，PEREZ-FONS L，ROBERTSON F P，et al.Functional characterization of long-chain prenyl diphosphate synthases from tomato[J].Biochemical Journal，2013，449(3)：729-740.

[2]DOYLE M，LI L，YU L，et al.Identification of a Mr=17,000 protein as the plastoquinone-binding protein in the cytochrome b6-f complex from spinach chloroplasts[J].Journal of Biological Chemistry，1989，264(3)：1387-1392.

[3]ERNSTER L，LEE I Y，NORLING B，et al.Studies with ubiquinone-depleted submitochondrial particles[J].European Journal of Biochemistry，1969，9(3)：299-310.

[4]JUN L，SAIKI R，TATSUMI K，et al.Identification and subcellular localization of two solanesyl diphosphate synthases from Arabidopsis thaliana[J].Plant and Cell Physiology，2004，45(12)：1882-1888.

[5]RODRIGUEZ-CONCEPCION M，BORONAT A.Elucidation of the methylerythritol phosphate pathway for isoprenoid biosynthesis in bacteria and plastids.A metabolic milestone achieved through genomics[J].Plant Physiology，2002，130(3)：1079-1089.

[6]DISCH A，HEMMERLIN A，BACH T，et al.Mevalonate-derived isopentenyl diphosphate is the biosynthetic precursor of ubiquinone prenyl side chain in tobacco BY-2 cells[J].Biochemical Journal，1998，331：615-621.

[7]FUKUSAKI E，TAKENO S，BAMBA T，et al.Biosynthetic pathway for the C45 polyprenol，solanesol，in tobacco[J].Bioscience，Biotechnology and Biochemistry，2004，68(9)：1988-1990.

[8]OKADA K，SUZUKI K，KAMIYA Y，et al.Polyprenyl diphosphate synthase essentially defines the length of the side chain of ubiquinone[J]. Biochimica Biophysica Acta，1996，1302(3)：217-223.

[9]OKADA K，KAMIYA Y，ZHU X，et al.Cloning of the sdsA gene encoding solanesyl diphosphate synthase from Rhodobacter capsulatus and its functional expression in Escherichia coli and Saccharomyces cerevisiae[J].Journal of Bacteriology，1997，179(19)：5992-5998.

[10]LIANG P H，KO T P，WANG A H.Structure，mechanism and function of prenyltransferases[J].European Journal of Biochemistry，2002，269(14)：3339-3354.

[11]OHARA K，SASAKI K，YAZAKI K.Two solanesyl diphosphate synthases with different subcellular localizations and their respective physiological roles in Oryza sativa[J].Journal of Experimental Botany，2010，61(10)：2683-2692.

[12]HIROOKA K，BAMBA T，FUKUSAKI E，et al.Cloning and kinetic characterization of Arabidopsis thaliana solanesyl diphosphate synthase[J].Biochemical Journal，2003，370：679-686.

[13]李红伟，郑晓珂，弓建红，等.独行菜和播娘蒿化学成分及药理作用研究进展[J].药物评价研究，2013(3)：235-240.

[14]ROSANO G L，CECCARELLI E A.Recombinant protein expression in Escherichia coli：advances and challenges[J].Frontiers in Microbiology，2014，5(4)：172.

Memo

Memo:
-
Last Update: 2016-05-03