The substitution of cysteine 17 of recombinant human G-CSF with alanine greatly enhanced its stability.
Human recombinant granulocyte-colony stimulating factor (rhG-CSF) has one free cysteine at position 17 and has two disulfide bridges (Cys36-Cys42 and Cys64-Cys74). The Cys17 of rhG-CSF was substituted with Gly, Ala, Ser, Ile, Tyr, Arg, and Pro, or deleted using site-directed mutagenesis in order to improve its thermostability. With the exception of Pro17-rhG-CSF, all mutant proteins retained biological activity which promotes the growth of mouse bone marrow cells in vitro. Among these mutant proteins, Ala17-rhG-CSF had more than 5 times higher stability than rhG-CSF. But Ser17-rhG-CSF had almost same stability as rhG-CSF and other mutant proteins had only lower stability.
Ishikawa
M
M
Pharmaceutical Laboratory, Kirin Brewery Co., Ltd., Gunma, Japan.
Iijima
H
H
Satake-Ishikawa
R
R
Tsumura
H
H
Iwamatsu
A
A
Kadoya
T
T
Shimada
Y
Y
Fukamachi
H
H
Kobayashi
K
K
Matsuki
S
S
D000409:Alanine / Q000737:chemistry* / Q000235:genetics; D000818:Animals; D001854:Bone Marrow Cells; D003545:Cysteine / Q000737:chemistry* / Q000235:genetics; D005260:Female; D016179:Granulocyte Colony-Stimulating Factor / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D006358:Hot Temperature; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D008807:Mice, Inbred BALB C; D016297:Mutagenesis, Site-Directed / Q000502:physiology; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D013329:Structure-Activity Relationship
D011994:Recombinant Proteins; D016179:Granulocyte Colony-Stimulating Factor; D003545:Cysteine; D000409:Alanine