Multiple-copy genes: production and modification of monomeric peptides from large multimeric fusion proteins.
A vector system has been designed for obtaining high yields of polypeptides synthesized in Escherichia coli. Multiple copies of a synthetic gene encoding the neuropeptide substance P (SP) (Arg-Pro-Lys-Pro-Gln-Gln-Phe-Phe-Gly-Leu-Met-NH2) have been linked and fused to the lacZ gene. Each copy of the SP gene was flanked by codons for methionine to create sites for cleavage by cyanogen bromide (CNBr). The isolated multimeric SP fusion protein was converted to monomers of SP analog, each containing a carboxyl-terminal homoserine lactone (Hse-lactone) residue (Arg-Pro-Lys-Pro-Gln-Gln-Phe-Phe-Gly-Leu-Hse-lactone), upon treatment with CNBr in formic acid. The Hse-lactone moiety was subjected to chemical modifications to produce an SP Hse amide. This method permits synthesis of peptide amide analogs and other peptide derivatives by combining recombinant DNA techniques and chemical methods.
Kempe
T
T
Kent
S B
SB
Chow
F
F
Peterson
S M
SM
Sundquist
W I
WI
L'Italien
J J
JJ
Harbrecht
D
D
Plunkett
D
D
DeLorbe
W J
WJ
D003001:Cloning, Molecular / Q000379:methods*; D003488:Cyanogen Bromide; D004274:DNA, Recombinant; D004926:Escherichia coli / Q000235:genetics; D005786:Gene Expression Regulation; D005822:Genetic Vectors; D006801:Humans; D008970:Molecular Weight; D010446:Peptide Fragments; D010957:Plasmids; D013373:Substance P / Q000235:genetics*; D001616:beta-Galactosidase / Q000235:genetics
D004274:DNA, Recombinant; D010446:Peptide Fragments; D013373:Substance P; D001616:beta-Galactosidase; D003488:Cyanogen Bromide