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Titolo:
Heterogeneous distribution of lysine 6-aminotransferase during cephamycin C biosynthesis in Streptomyces clavuligerus demonstrated using green fluorescent protein as a reporter
Autore:
Khetan, A; Hu, WS; Sherman, DH;
Indirizzi:
Univ Minnesota, Inst Technol, Dept Microbiol & Biol Proc, Minneapolis, MN 55455 USA Univ Minnesota Minneapolis MN USA 55455 l Proc, Minneapolis, MN 55455 USA Univ Minnesota, Dept Chem Engn & Mat Sci, Minneapolis, MN 55455 USA Univ Minnesota Minneapolis MN USA 55455 at Sci, Minneapolis, MN 55455 USA
Titolo Testata:
MICROBIOLOGY-UK
, volume: 146, anno: 2000,
parte:, 8
pagine: 1869 - 1880
SICI:
1350-0872(200008)146:<1869:HDOL6D>2.0.ZU;2-7
Fonte:
ISI
Lingua:
ENG
Soggetto:
GENE-EXPRESSION; EPSILON-AMINOTRANSFERASE; BACILLUS-SUBTILIS; QUANTITATIVE CHARACTERIZATION; CEPHALOSPORIN BIOSYNTHESIS; COELICOLOR A3(2); LOCALIZATION; SPORULATION; CELLS; SYNTHETASE;
Keywords:
Streptomyces clavuligerus; lysine 6-aminotransferase; cephamycin biosynthesis; heterogeneity; GFP;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
49
Recensione:
Indirizzi per estratti:
Indirizzo: Sherman, DH Univ Minnesota, Inst Technol, Dept Microbiol & Biol Proc, Box 196,1460 Mayo Mem Bldg,420 Delaware St SE, Minneapolis, MN 55455 USA Univ Minnesota Box 196,1460 Mayo Mem Bldg,420 Delaware St SE Minneapolis MN USA 55455
Citazione:
A. Khetan et al., "Heterogeneous distribution of lysine 6-aminotransferase during cephamycin C biosynthesis in Streptomyces clavuligerus demonstrated using green fluorescent protein as a reporter", MICROBIO-UK, 146, 2000, pp. 1869-1880

Abstract

The cellular distribution of the cephamycin biosynthetic enzyme lysine 6-aminotransferase (LAT) has been studied in Streptomyces clavuligerus hyphae by confocal microscopy using the S65T mutant of green fluorescent protein (GFP) as a reporter, LAT mediates the first committed step in the biosynthesis Of the secondary metabolite cephamycin C by S. clavuligerus. The enzymicactivity of LAT varies with time during the growth of S, clavuligerus in liquid medium. To investigate if this temporal variation occurs uniformly amongst all hyphae, S, clavuligerus was transformed with a plasmid containingthe LAT-encoding gene translationally fused to the GFP-encoding gene. The LAT-GFP fusion product displayed fluorescence spectral characteristics of GFP, and showed similar temporal characteristics of LAT activity compared tothe wild-type strain of S, clavuligerus, The transformed strain exhibited a heterogeneous distribution of fluorescence in mycelia grown in liquid cultures. This distribution varied significantly as the batch progressed: onlya fraction of the mycelia fluoresced in the early growth phase, whereas nearly all hyphae fluoresced by the late growth phase. Thereafter, a non-uniform distribution of fluorescence was again observed in the declining growthphase. A large fraction of the non-fluorescent cells in the declining growth phase were found to be non-viable. Observations of S, clavuligerus colonies grown on solid agar also showed variation of LAT-GFP expression at different stages of growth. These observations in the solid phase can be explained in terms of nutrient deprivation and signalling molecules. The results suggest that physiological differentiation of S, clavuligerus mycelia leading to cephamycin C biosynthesis is both temporally and spatially distributed. The findings also revealed that the observed heterogeneity was independent of the position of individual cell compartments within the hypha, The potential of GFP as a reporter for the quantitative study of cephamycin biosynthesis at the cellular level has also been demonstrated.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 30/09/20 alle ore 22:44:26