Molecular cloning, expression in Escherichia coli and production of bioactive homogeneous recombinant human granulocyte and macrophage colony stimulating factor.
Human granulocyte and macrophage colony stimulating factor (hGM-CSF) is a glycoprotein that activates and enhances the differentiation and survival of neutrophils, eosinophils and macrophages, which play a key role in the innate immune response. Here we describe the construction of the hGM-CSF encoding gene, cloning, expression in Escherichia coli, purification of recombinant hGM-CSF, N-terminal amino acid sequencing, and biological activity assay using TF-1 cells. The results presented show that the combination of experimental strategies employed to obtain recombinant hGM-CSF can yield biologically active protein, and may be useful to scaling-up production of biosimilar protein.
Schwanke
Raquel Cristina
RC
Centro de Pesquisas em Biologia Molecular e Funcional, Instituto Nacional de Ciência e Tecnologia em Tuberculose, Pontifícia Universidade Católica do Rio Grande do Sul (PUCRS), Porto Alegre-RS 90619-900, Brazil.
Renard
Gaby
G
Chies
Jocelei Maria
JM
Campos
Maria Martha
MM
Batista
Eraldo Luiz
EL
Santos
Diógenes Santiago
DS
Basso
Luiz Augusto
LA
D000595:Amino Acid Sequence; D000818:Animals; D045744:Cell Line, Tumor; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D004926:Escherichia coli / Q000166:cytology / Q000235:genetics*; D015870:Gene Expression; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D006801:Humans; D002479:Inclusion Bodies / Q000235:genetics; D008969:Molecular Sequence Data; D011994:Recombinant Proteins
D018076:DNA, Complementary; D011994:Recombinant Proteins; D016178:Granulocyte-Macrophage Colony-Stimulating Factor