Expression of the carboxypeptidase T gene from Thermoactinomyces vulgaris in stable protoplast type L-forms of Proteus mirabilis.
The structural gene of the carboxypeptidase T (cpt) was successfully expressed in cell wall-less L-form cells of Proteus mirabilis. The DNA sequence encoding the PhoA leader peptide was fused with a truncated cpt gene encoding the mature enzyme. The modified gene in a pUC-based kanamycin resistance vector under the control of the lac promoter was transformed into L-form cells of P. mirabilis. They were able to produce the recombinant CpT both as a secretory and as a cell-bound insoluble form. The co-secretory processing of the PhoA leader peptide was quite efficient. The yield of the secreted CpT was not less than 20 mg l-1 and should be improvable.
Bushueva
A M
AM
Laboratory of Protein Chemistry, Institute of Microbial Genetics (GNII Genetika), Moscow, Russia.
Shevelev
A B
AB
Gumpert
J
J
Chestukhina
G G
GG
Serkina
A V
AV
Hoischen
C
C
Matz
M V
MV
Kuryatova
M V
MV
Stepanov
V M
VM
D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D001483:Base Sequence; D002268:Carboxypeptidases / Q000235:genetics*; D007740:L Forms / Q000235:genetics*; D008848:Micromonosporaceae / Q000235:genetics*; D008969:Molecular Sequence Data; D011513:Proteus mirabilis / Q000235:genetics*; D011994:Recombinant Proteins / Q000378:metabolism
D001426:Bacterial Proteins; D011994:Recombinant Proteins; D002268:Carboxypeptidases; C076614:CPT protein, Thermoactinomyces vulgaris