[Deletion mutants of human granulocyte-macrophage colony-stimulating factor].
To study the structure-function relationship of the human granulocyte-macrophage colony-stimulating factor (GM-CSF), genes were constructed that encode its three deletion mutants: D1, a mutant with the deletion of six amino acid residues (37-42) some of which are a part of a beta-structural region; D2, a mutant with the deletion of the unstructured six-aa sequence of a loop (45-50); and D3, a mutant with the deletion of 14 aa residues (37-50) corresponding to the A-B loop and encoded by the second exon of the gmcsf gene. The expression products of these genes in E. coli were accumulated in a fraction of insoluble proteins. The secondary structures of the mutant proteins were similar to that of the full-size GM-CSF, but the biological activity of the deletion mutants was 130 times lower than that of the GM-CSF: they stimulated the proliferation of the TF-1 cell line at 3 ng/ml concentration. The resulting proteins displayed antagonistic properties toward the full-size GM-CSF, with the inhibition degree of its colony-stimulating activity being 27%. A decrease in the mutant activity in the row D2 > D1 > D3 implies the importance of the conserved hydrophobic residues involved in the formation of the beta-structure for the formation of the GM-CSF functional conformation.
Petrovskaia
L E
LE
Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, ul. Miklukho-Maklaya 16/10, GSP Moscow, 117997 Russia. lpetr@mail.ibch.ru
Kriukova
E A
EA
Kaiushin
A L
AL
Rodina
A V
AV
Moskaleva
E Iu
EIu
Korobko
V G
VG
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005813:Genes, Synthetic / Q000235:genetics; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000235:genetics* / Q000494:pharmacology; D006801:Humans; D008969:Molecular Sequence Data; D011994:Recombinant Proteins; D015183:Restriction Mapping; D017384:Sequence Deletion
D011994:Recombinant Proteins; D016178:Granulocyte-Macrophage Colony-Stimulating Factor