[Construction of BPI23-Fc gamma 1 recombinant protein prokaryotic expression vector and the expression and biological assessment of BPI23-Fc gamma 1 recombinant protein].
OBJECTIVE
To construct pBV-BPI600-Fc gamma 1(700) recombinant expression vector, to transform into Escherichia coli DH5 alpha, and to induce the expression of BPI23-Fc gamma 1 anti-bacterial recombinant protein.


METHODS
Genes which encode BPI23 and Fc gamma 1 were amplified by RT-PCR from mRNA that was extracted from HL-60 cell and normal human leukocytes; Recombinant cloning vector and recombinant expression vector were constructed. pBV-BPI600-Fc gamma 1(700) recombinant expression vector was transformed into the competent Escherichia coli DH5 alpha and BPI23-Fc gamma 1 recombinant protein was expressed by temperature induced method.


RESULTS
(1) Expected amplified products BPI600 bp and Fc gamma 1(700) bp were obtained by RT-PCR. (2) pUC18-BPI180, pUC18-BPI420, and pUC18-Fc gamma 1(700) recombinant cloning vector were successfully constructed, and sequences were identical with the reported ones. (3) pBV-BPI600-Fc gamma 1(700) recombinant expression vector was successfully constructed, and results of the enzyme digestion analysis were identical with expected ones. (4) pBV-BPI600-Fc gamma 1(700) recombinant expression vector was transformed into the competent Escherichia coli DH5 alpha and BPI23-Fc gamma 1 recombinant protein was expressed by temperature-induced method, and its expression level was accounted for 20% of total bacterial proteins. (5) The renatured BPI23-Fc gamma 1 recombinant protein had anti-bacterial activity and biological functions of complement fixation, opsonization.


CONCLUSIONS
pBV-BPI600-Fc gamma 1(700) recombinant expression vector was successfully constructed, and BPI23-Fc gamma 1 recombinant protein with BPI and IgGFc double biological activity was expressed in Escherichia coli.
An
Y
Y
Department of Microbiology and Immunology, Capital University of Medical Sciences, Beijing 100054, China. anyunq@cpums.edu.cn
Ke
Y
Y
Yang
G
G
D023181:Antimicrobial Cationic Peptides; D001798:Blood Proteins / Q000096:biosynthesis* / Q000235:genetics; D002478:Cells, Cultured; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005822:Genetic Vectors; D018922:HL-60 Cells; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000096:biosynthesis* / Q000235:genetics; D007074:Immunoglobulin G / Q000096:biosynthesis* / Q000235:genetics; D007962:Leukocytes / Q000166:cytology; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction
D023181:Antimicrobial Cationic Peptides; D001798:Blood Proteins; D007141:Immunoglobulin Fc Fragments; D007074:Immunoglobulin G; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; C021623:bactericidal permeability increasing protein