Site-specific mutagenesis of the human interleukin-1 beta gene: structure-function analysis of the cysteine residues.
Human interleukin-1 beta (IL-1 beta) has two cysteines located at amino acid residues 8 and 71 of the mature protein consisting 153 amino acids. To clarify the role of these characteristic cysteine residues in IL-1 beta, at first, an expression plasmid for site-specific mutagenesis has been constructed by inserting the ori and intergenic region of phage f1 into the IL-1 beta expression vector. The plasmid can be used not only for isolation of the modified IL-1 beta gene but for expression of the mutant protein in Escherichia coli. Using this plasmid, each of the cysteine codons in IL-1 beta gene was changed to serine or alanine codon, or deleted. The modified IL-1 beta showed that the two cysteine residues in IL-1 beta are not essential for biological activity but not to be eliminated for the maintenance of the functional structure of IL-1 beta.
Kamogashira
T
T
Laboratories of Cellular Technology, Otsuka Pharmaceutical Co., Ltd., Tokushima, Japan.
Masui
Y
Y
Ohmoto
Y
Y
Hirato
T
T
Nagamura
K
K
Mizuno
K
K
Hong
Y M
YM
Kikumoto
Y
Y
Nakai
S
S
Hirai
Y
Y
D003062:Codon; D003545:Cysteine; D004262:DNA Restriction Enzymes; D004274:DNA, Recombinant; D004277:DNA, Single-Stranded / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D007375:Interleukin-1 / Q000235:genetics* / Q000494:pharmacology; D008545:Melanoma / Q000473:pathology; D008970:Molecular Weight; D009154:Mutation; D010957:Plasmids; D013329:Structure-Activity Relationship; D014407:Tumor Cells, Cultured
D003062:Codon; D004274:DNA, Recombinant; D004277:DNA, Single-Stranded; D007375:Interleukin-1; D004262:DNA Restriction Enzymes; D003545:Cysteine