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Titolo:
PECTIN METHYL ESTERASE FROM ORANGE FRUIT - CHARACTERIZATION AND LOCALIZATION BY IN-SITU HYBRIDIZATION AND IMMUNOHISTOCHEMISTRY
Autore:
CHRISTENSEN TMIE; NIELSEN JE; KREIBERG JD; RASMUSSEN P; MIKKELSEN JD;
Indirizzi:
DANISCO BIOTECHNOL,LANGEBROGADE 1 DK-1001 COPENHAGEN K DENMARK
Titolo Testata:
Planta
fascicolo: 4, volume: 206, anno: 1998,
pagine: 493 - 503
SICI:
0032-0935(1998)206:4<493:PMEFOF>2.0.ZU;2-W
Fonte:
ISI
Lingua:
ENG
Soggetto:
PHASEOLUS-VULGARIS L; PARTIAL-PURIFICATION; CELL-WALLS; METHYLESTERASE; PECTINESTERASE; TOMATO; EXPRESSION; TISSUES; PROTEINS; FLAX;
Keywords:
CDNA CLONE; CELL WALL; CITRUS (FRUIT, PECTIN); FRUIT RIPENING; PECTIN DEGRADATION; PECTIN METHYL ESTERASE (PURIFICATION, LOCALIZATION);
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Science Citation Index Expanded
Citazioni:
38
Recensione:
Indirizzi per estratti:
Citazione:
T.M.I.E. Christensen et al., "PECTIN METHYL ESTERASE FROM ORANGE FRUIT - CHARACTERIZATION AND LOCALIZATION BY IN-SITU HYBRIDIZATION AND IMMUNOHISTOCHEMISTRY", Planta, 206(4), 1998, pp. 493-503

Abstract

Pectin methyl esterase (PME) from orange (Citrus sinensis L.) fruit peels has been purified by ammonium sulphate precipitation, and ion-exchange and gel-filtration chromatography. Characterization of the enzyme revealed a 36-kDa protein with an isoelectric point > 9, a pH optimum at 7 and temperature optimum at 50 degrees C. The substrate specificity and kinetic experiments showed that the affinity of PME for pectinwas highly dependent on the degree of esterification (DE) of the pectin, with K-m values of 0.7 mg ml(-1) for pectin with a DE of 70% and 17 mg ml(-1) for pectin with;a DE of 25%. The sequences of the NH2-terminal end of digested peptides from the mature protein were obtained. ADNA fragment of 501 bp was cloned by polymerase chain reaction amplification using degenerate primers and was further used for screening ofa cDNA library. Two cDNA clones were isolated encoding PMEs of 584 amino acids and 362 amino acids, respectively, including a putative signal peptide. The deduced amino acid sequence showed full identity to the sequenced peptides. Polyclonal antibodies raised against orange peelPME were used for immunohistochemistry. The main localization of PMEswas in the outer cell layers of the juice vesicles, in the outer celllayers of the lamellae between the segments and in the inner cell layers of the albedo in the peel. In-situ hybridization showed that the mRNA is very abundant in the fruit and was found in the same cell layers as the native enzyme. A very intensive staining for PME mRNA was also seen in the core and in the flavedo close to the oil glands.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 31/03/20 alle ore 10:12:18