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Titolo:
High-level expression, purification, and some properties of a recombinant cephalosporin-C deacetylase
Autore:
Takimoto, A; Yagi, S; Mitsushima, K;
Indirizzi:
Shionogi & Co Ltd, Proc R&D Labs, Bioproc Program, Amagasaki, Hyogo 6600813, Japan Shionogi & Co Ltd Amagasaki Hyogo Japan 6600813 aki, Hyogo 6600813, Japan
Titolo Testata:
JOURNAL OF BIOSCIENCE AND BIOENGINEERING
fascicolo: 4, volume: 87, anno: 1999,
pagine: 456 - 462
SICI:
1389-1723(199904)87:4<456:HEPASP>2.0.ZU;2-I
Fonte:
ISI
Lingua:
ENG
Soggetto:
ESCHERICHIA-COLI; BACILLUS-SUBTILIS; GROWTH-HORMONE; SEQUENCES; PROTEINS;
Keywords:
cephalosporin-C deacetylase; Bacillus subtilis; Escherichia coli; production; purification; properties;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Agriculture,Biology & Environmental Sciences
Life Sciences
Citazioni:
20
Recensione:
Indirizzi per estratti:
Indirizzo: Takimoto, A Shionogi & Co Ltd, Proc R&D Labs, Bioproc Program, 1-3 Kuise Terajima 2 Chome, Amagasaki, Hyogo 6600813, Japan Shionogi & Co Ltd 1-3 Kuise Terajima 2 Chome Amagasaki Hyogo Japan 6600813
Citazione:
A. Takimoto et al., "High-level expression, purification, and some properties of a recombinant cephalosporin-C deacetylase", J BIOSCI BI, 87(4), 1999, pp. 456-462

Abstract

To maximize the expression of the cephalosporin-C deacetylase (CAH) gene isolated from Bacillus subtilis SHS 0133 in Escherichia coli, a series of expression plasmids was constructed with various spacings between the Shine-Dalgarno sequence and the ATG initiation codon. As the most efficient expression plasmid, we selected pCAH431, which has the trp promoter, a replication origin derived from pAT153, and a spacing of 13 nucleotides. E. coli JM103 with pCAH431 produced 4.9 g of CAH per liter on cultivation at 37 degreesC for 20 h in a 30-l jar fermenter. Since the amount of CAH reached about 70% of the total protein h the soluble fraction of the cells, and CAB was recovered from the cell extracts in an active form, the CAH was purified easily to homogeneity by only one column chromatography step, Twenty grams of 7-aminocephalosporanic acid was completely converted to deacetyl-7-aminocephalosporanic acid, a starting material for cefcapene pivoxil hydrochloride,by 12 mg of the purified enzyme without significant appearance of by-products. Thus, our expression and purification system has made the industrial production of CAH possible.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 28/09/20 alle ore 13:03:43