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Titolo:
THE ALKB MONOOXYGENASE OF PSEUDOMONAS-OLEOVORANS - SYNTHESIS, STABILITY AND LEVEL IN RECOMBINANT ESCHERICHIA-COLI AND THE NATIVE HOST
Autore:
STAIJEN IE; HATZIMANIKATIS V; WITHOLT B;
Indirizzi:
ETH HONGGERBERG,ETH ZURICH,INST BIOTECHNOL CH-8093 ZURICH SWITZERLAND ETH HONGGERBERG,ETH ZURICH,INST BIOTECHNOL CH-8093 ZURICH SWITZERLAND
Titolo Testata:
European journal of biochemistry
fascicolo: 2, volume: 244, anno: 1997,
pagine: 462 - 470
SICI:
0014-2956(1997)244:2<462:TAMOP->2.0.ZU;2-2
Fonte:
ISI
Lingua:
ENG
Soggetto:
ENZYMATIC OMEGA-OXIDATION; ALKANE HYDROXYLASE; RUBREDOXIN REDUCTASE; PROTEINS; SYSTEM; EXPRESSION; PLASMIDS; MUTANTS; OCTANE; PUTIDA;
Keywords:
ALKANE MONOOXYGENASE; MEMBRANE PROTEIN; SYNTHESIS; STABILITY; PSEUDOMONAS OLEOVORANS;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Science Citation Index Expanded
Citazioni:
51
Recensione:
Indirizzi per estratti:
Citazione:
I.E. Staijen et al., "THE ALKB MONOOXYGENASE OF PSEUDOMONAS-OLEOVORANS - SYNTHESIS, STABILITY AND LEVEL IN RECOMBINANT ESCHERICHIA-COLI AND THE NATIVE HOST", European journal of biochemistry, 244(2), 1997, pp. 462-470

Abstract

We have studied the synthesis and stability of the monooxygenase AlkBof Pseudomonas oleovorans in its natural host and in recombinant Escherichia coli. Three strains were investigated: the prototype strain P.oleovorans and the E. coli alk(+) recombinants HB101 (pGEc47) and W3110 (pGEc47). Plasmid pGEc47 allows regulated expression of alkB and synthesis of active AlkB in E. coli. The E. coli strains were selected because E. coli HB101 (pGEc47) produces similar amounts of AlkB as P. oleovorans (1.5-2% of total cell protein), whereas E. coli W3110 (pGEc47) is able to make substantially (about fivefold) more AlkB. The AlkB synthesis and degradation rates in batch cultures of the three strainswere determined by means of isotopic-labeling and immunological techniques. The mean specific AlkB synthesis rates in P. oleovorans, E. coli HB101 (pGEc47) and E. coli W3110 (pGEc47) were approximately 7, 12.5and 45 mu g . mg . protein(-1) . h(-1), respectively. The half-lives of AlkB were estimated to be 80, 3 and 15 for P. oleovorans, E. coli HB101 (pGEc47) and E. coli W3110 (pGEc47), respectively. Thus, the intracellular AlkB level in each of the three strains was the result of their AlkB synthesis and degradation rates. The AlkB level during batch growth was modelled by means of experimentally derived parameters for AlkB synthesis and degradation, and showed good agreement with AlkB levels determined by means of immunoblotting in all strains investigated.

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Documento generato il 29/11/20 alle ore 00:22:50