|Cameron, Randall - Randy|
|Kim, Yan - Former Ars Employee|
|Tzen, Jason - National Chung-hsing University|
Submitted to: Food Hydrocolloids
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/15/2015
Publication Date: 2/11/2015
Citation: Cameron, R.G., Kim, Y., Galant, A.L., Luzio, G.A., Tzen, J. 2015. Pectin Homogalacturonans: Nanostructural Characterization of Methylesterified Domains. Food Hydrocolloids Journal. 47:184-190. Interpretive Summary: The growth of plant cells is mediated by both a complex set of molecules and a series of events resulting in cell wall expansion. The dominant molecular species associated with plant cell walls are cellulose, hemicellulose and pectin. Recently the role of pectin in cell wall organization and functionality has received considerable attention. The structural features of pectin most often discussed are those related to both the net charge and the distribution of charge along its largely linear molecules. These are also the structural features that are frequently listed as the basis for pectin’s technological and economical attributes, which focus on its ability to crosslink with divalent cations to alter the texture of foods and the rheology of water as well as to bind metals. The structural complexity of pectin is well established and necessitates a statistical approach for describing a population of pectin molecules. Because of its functional importance more attention has been focused on characterizing the charged, non methylesterified domains (NMD) of the dominant homogalacturonan region (HG) and the enzyme mode of action that produces it rather than on the neutral, methylesterified domains. More recently some attention has been turned towards these more neutral, uncharged, methylesterified domains. These methyl protected domains (MPD) also have both biological and technological significance because they are capable of participating in hydrogen bonding and hydrophobic interactions between adjacent pectin molecules and are resistant to depolymerization by fungal endo polygalacturonases which require some minimal number of contiguous non methylesterified galacturonic acid (GalA) units for hydrolysis to occur. Here we present a method to enzymatically digest the HG NMDs, leaving only the MPDs intact. Following demethylesterification the resulting GalA oligomers were separated and chromatographically visualized allowing for estimation of the mass and concentration of individual oligomers. Subsequently the parameters “average number of MPDs per molecule”, “sum of GalAs” in these MPDs per molecule, “average MPD size” and the “number of average sized MPDs per molecule” are introduced.
Technical Abstract: Functionality of pectic hydrocolloids is largely dependent on the two major domains commonly found in their homogalacturonan (HG) regions, i.e., methylester protected domains (MPDs)and non methylesterified domains (NMDs). MPDs can participate in hydrogen bonding and hydrophobic interactions but unlike the NMDs they are incapable of calcium mediated cross-linking. MPDs also are inaccessible to endo polygalacturonase attack while NMDs are susceptible. We isolated and characterized MPDs contained within homogalacturonans that had been demethylesterified either by plant pectin methylesterases or chemically by base saponification. Molecular weight (~ 9500 - 14,000 Da) and degree of methylesterification (DM; ~ 63% - 94%) of remnant MPDs were measured. Subsequently the MPDs were base saponified at 4 °C. The resulting demethylesterified oligomers were separated, visualized and quantified. Population parameters estimated were the average number of MPDs per pectin molecule, the sum of galacturonic acid units in MPDs per molecule, the average MPD size and the number of "average" size MPDs per molecule. Analysis of variance and Tukey's Multiple Comparison Test indicated significant differences in MPDs based on DM and the method of demethylesterification. Pearson's correlation coefficients were calculated for MPD parameters, previously published structural data from NMDs and rheological properties of the demethylesterified homogalacturonans from the enzyme treatments. Significant correlations were observed for DM, absolute degree of blockiness, G', NMD and MPD parameters. When coupled with methods to characterize NMDs these techniques provide an opportunity to statistically describe both highly charged and more neutral domains in a population of pectin molecules.