Mutagenesis of human granulocyte colony stimulating factor.
To define the structure-function relationship, we have made a number of mutants of human granulocyte colony-stimulating factor (hG-CSF) by in vitro mutagenesis. The results indicate that most of the mutations located in the internal and C-terminal regions of the molecule abolished the activity, whereas the mutants without N-terminal 4, 5, 7, or 11 amino acids retained the activity. N-terminal amino acids were also altered by cassette mutagenesis using a synthetic oligonucleotide mixture. Among them, KW2228, in which Thr-1, Leu-3, Gly-4, Pro-5 and Cys-17 were respectively substituted with Ala, Thr, Tyr, Arg and Ser, showed more potent granulopoietic activity than that of intact hG-CSF both in vitro and in vivo.
Kuga
T
T
Tokyo Research Laboratories, Kyowa Hakko Kogyo Co. Ltd., Japan.
Komatsu
Y
Y
Yamasaki
M
M
Sekine
S
S
Miyaji
H
H
Nishi
T
T
Sato
M
M
Yokoo
Y
Y
Asano
M
M
Okabe
M
M
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001681:Biological Assay; D001854:Bone Marrow Cells; D003062:Codon; D003115:Colony-Stimulating Factors / Q000235:genetics / Q000502:physiology*; D004247:DNA / Q000235:genetics; D004262:DNA Restriction Enzymes; D004274:DNA, Recombinant; D004926:Escherichia coli / Q000235:genetics; D016179:Granulocyte Colony-Stimulating Factor; D006098:Granulocytes / Q000166:cytology; D006410:Hematopoiesis; D006801:Humans; D008297:Male; D051379:Mice; D008809:Mice, Inbred C3H; D008969:Molecular Sequence Data; D009154:Mutation; D013329:Structure-Activity Relationship
D003062:Codon; D003115:Colony-Stimulating Factors; D004274:DNA, Recombinant; D016179:Granulocyte Colony-Stimulating Factor; D004247:DNA; D004262:DNA Restriction Enzymes