[Cloning and eukaryotic expression of human lipocalin-type prostaglandin D synthase in Pichia Pastoris].
OBJECTIVE
To express human testis Lipocalin-type prostaglandin D synthase in Pichia Pastoris for further research on biological function and clinical applications.


METHODS
Human testis L-PGDS gene coding region was amplified from plasmid pGEX-2T/htL-PGDS by PCR with a deletion of the signal peptide sequence. The DNA fragment was inserted into pPIC9 to construct yeast expression plasmid followed by transformation of the yeast GS115 strain with electroporation. The recombinant his-tag protein was induced to express by methanol.


RESULTS
The sequence of the amplified DNA fragment was identical to that of human testis L-PGDS previously reported. The recombinant protein was found with a molecular mass of 27,000 on SDS-PAGE, which was identical to that of native L-PGDS.


CONCLUSIONS
Secretory expression of human L-PGDS was obtained in Pichia Pastoris.
Department of Immunology and Microbiology, Nanjing Medical University, Nanjing, Jiangsu 210029, China.
Yun
Y
Gao
Yu-Feng
YF
Huang
Xin-Yi
XY
Xia
Bai-Kun
BK
Ma
D003001:Cloning, Molecular; D015870:Gene Expression; D006801:Humans; D019746:Intramolecular Oxidoreductases / Q000096:biosynthesis* / Q000235:genetics; D054834:Lipocalins; D008297:Male; D010843:Pichia / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism
D054834:Lipocalins; D011994:Recombinant Proteins; D019746:Intramolecular Oxidoreductases; C022466:prostaglandin R2 D-isomerase