title: Pectin methylesterase in Citrus bergamia R.: purification, biochemical characterisation and sequence of the exon related to the enzyme active site.
abstract: Three forms of pectin methylesterase (PME) were purified, from bergamot fruit (Citrus bergamia R.), to homogeneity by ion-exchange and affinity chromatography. The isoforms, named PME I, PME II and PME III, according their elution order on a heparin-sepharose column, were characterized for their relative molecular mass, activity kinetic parameters and thermostability. The molecular mass was estimated to be 42kDa for the three forms, and the apparent Km values for citrus pectin were 0.9mg/ml for PME I and 0.5mg/ml for PME II and PME III. The optimum pH values lie within the range 6.5-9.0, depending on salt concentration. Thermal behaviours of the three PME isoforms were studied in a temperature range from 65°C to 80°C with the less abundant PME I isoform showing a higher heat resistance. Moreover, the complete exon 2 sequence of PME gene was acquired (GenBank accession no. DQ458770) using a PCR-based approach on well-known Citrus genomic DNA present in the NCBI database.
authors.affiliation: Istituto di Chimica Biomolecolare (ICB), Consiglio Nazionale delle Ricerche (CNR), Via Campi Flegrei, 34-80078 Pozzuoli (NA), Italy. Electronic address: blaratta@icmib.na.cnr.it.
authors.forename: Bruna
authors.initials: B
authors.lastname: Laratta
authors.affiliation: Istituto di Genetica Vegetale (IGV), Consiglio Nazionale delle Ricerche (CNR), Via Università, 133-80055 Portici (NA), Italy.
authors.forename: Luigi
authors.initials: L
authors.lastname: De Masi
authors.affiliation: Stazione Sperimentale per le Industrie delle Essenze e dei Derivati Agrumari, Via Gen. Tommasini, 2-89127 Reggio Calabria, Italy.
authors.forename: Paola
authors.initials: P
authors.lastname: Minasi
authors.affiliation: Dipartimento di Biochimica e Biofisica "F. Cedrangolo", Seconda Università degli Studi di Napoli, Via S. M. di Costantinopoli, 16-80138 Napoli, Italy.
authors.forename: Alfonso
authors.initials: A
authors.lastname: Giovane
keywords: Characterization; Citrus bergamia; Gene sequence; Pectin methylesterase; Purification; Thermal stability