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Titolo:
POLYKETIDE SYNTHASE ACYL CARRIER PROTEINS FROM STREPTOMYCES - EXPRESSION IN ESCHERICHIA-COLI, PURIFICATION AND PARTIAL CHARACTERIZATION
Autore:
CROSBY J; SHERMAN DH; BIBB MJ; REVILL WP; HOPWOOD DA; SIMPSON TJ;
Indirizzi:
UNIV BRISTOL,SCH CHEM,CANTOCKS CLOSE BRISTOL BS8 1TS AVON ENGLAND UNIV BRISTOL,SCH CHEM BRISTOL BS8 1TS AVON ENGLAND JOHN INNES INST,JOHN INNES CTR NORWICH NR4 7UH NORFOLK ENGLAND
Titolo Testata:
Biochimica et biophysica acta. Protein structure and molecular enzymology
fascicolo: 1, volume: 1251, anno: 1995,
pagine: 32 - 42
SICI:
0167-4838(1995)1251:1<32:PSACPF>2.0.ZU;2-H
Fonte:
ISI
Lingua:
ENG
Soggetto:
FATTY-ACID BIOSYNTHESIS; COELICOLOR A3(2); SACCHAROPOLYSPORA-ERYTHRAEA; TETRACENOMYCIN-C; GENE-CLUSTER; ANTIBIOTIC BIOSYNTHESIS; ENGINEERED BIOSYNTHESIS; NUCLEOTIDE-SEQUENCE; GLAUCESCENS; ACTINORHODIN;
Keywords:
ACYL CARRIER PROTEIN; POLYKERIDE SYNTHASE; 4-PHOSPHOPANTETHEINE; ELECTROSPRAY MASS SPECTROMETRY; MALONYL TRANSFERASE; STREPTOMYCES;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Science Citation Index Expanded
Science Citation Index Expanded
Citazioni:
50
Recensione:
Indirizzi per estratti:
Citazione:
J. Crosby et al., "POLYKETIDE SYNTHASE ACYL CARRIER PROTEINS FROM STREPTOMYCES - EXPRESSION IN ESCHERICHIA-COLI, PURIFICATION AND PARTIAL CHARACTERIZATION", Biochimica et biophysica acta. Protein structure and molecular enzymology, 1251(1), 1995, pp. 32-42

Abstract

Acyl carrier proteins (ACPs) of the type II polyketide synthases for the aromatic antibiotics actinorhodin, granaticin, frenolicin and oxytetracycline were expressed in Escherichia coil downstream of an inducible phage T7 promoter. For the act and oetc genes, several of the first eight codons were changed to synonymous codons used in highly expressed E. coil genes. Correlated with these changes, the amounts of the act and otc ACPs purified from the recombinant E. coli cultures were anorder of magnitude greater than for the gra and fren ACPs expressed from the unmodified genes. Electrospray mass spectrometry (ESMS) of thepurified proteins confirmed their calculated M(r) based on the DNA sequences while also revealing that, in the act and gra ACP samples, some 2% and 30% of the holo-form of the protein was present (i.e., carrying the 4'-phosphopantetheine prosthetic group), with the remainder(and100% of the otc and fren samples) being in the apo-form. Increasing incubation time post heat induction led to an increase in act holo-ACP. The recombinant act and gra ACPs could function in vitro as substrates for an S. coelicolor malonyl CoA:ACP acyl transferase, as measured by the coupling of a labelled malonyl unit to the ACP; their quantitative abilities to do so correlated with the proportions of deduced holo form in the two samples.

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Documento generato il 27/11/20 alle ore 02:25:54