title: Enzymatic modification of a model homogalacturonan with the thermally tolerant pectin methylesterase from Citrus: 1. Nanostructural characterization, enzyme mode of action, and effect of pH.
abstract: Methyl ester distribution in pectin homogalacturonan has a major influence on functionality. Enzymatic engineering of the pectin nanostructure for tailoring functionality can expand the role of pectin as a food-formulating agent and the use of in situ modification in prepared foods. We report on the mode of action of a unique citrus thermally tolerant pectin methylesterase (TT-PME) and the nanostructural modifications that it produces. The enzyme was used to produce a controlled demethylesterification series from a model homogalacturonan. Oligogalacturonides released from the resulting demethylesterified blocks introduced by TT-PME using a limited endopolygalacturonase digestion were separated and quantified by high-pressure anion-exchange chromatography (HPAEC) coupled to an evaporative light-scattering detector (ELSD). The results were consistent with the predictions of a numerical simulation, which assumed a multiple-attack mechanism and a degree of processivity ∼10, at both pH 4.5 and 7.5. The average demethylesterified block size (0.6-2.8 nm) and number of average-sized blocks per molecule (0.8-1.9) differed, depending upon pH of the enzyme treatment. The mode of action of this enzyme and consequent nanostructural modifications of pectin differ from a previously characterized citrus salt-independent pectin methylesterase (SI-PME).
authors.affiliation: Citrus and Subtropical Products Laboratory, Agricultural Research Service, United States Department of Agriculture, 600 Avenue S., Northwest, Winter Haven, Florida 33881, United States. randall.cameron@ars.usda.gov
authors.forename: Randall G
authors.initials: RG
authors.lastname: Cameron
authors.forename: Gary A
authors.initials: GA
authors.lastname: Luzio
authors.forename: Prasanna
authors.initials: P
authors.lastname: Vasu
authors.forename: Brett J
authors.initials: BJ
authors.lastname: Savary
authors.forename: Martin A K
authors.initials: MA
authors.lastname: Williams
mesh_terms: D055162:Biocatalysis; D002265:Carboxylic Ester Hydrolases / Q000737:chemistry*; D002957:Citrus / Q000737:chemistry / Q000201:enzymology*; D004795:Enzyme Stability; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D008956:Models, Chemical; D008962:Models, Theoretical; D015394:Molecular Structure; D010368:Pectins / Q000737:chemistry*; D010940:Plant Proteins / Q000737:chemistry*
chemical_list: D010940:Plant Proteins; D010368:Pectins; D002265:Carboxylic Ester Hydrolases; C022323:pectinesterase; C003181:polygalacturonic acid