Activity of an atypical Arabidopsis thaliana pectin methylesterase
- 502 Downloads
An Arabidopsis thaliana pectin methylesterase that was not predicted to contain any signaling sequence was produced in E. coli and purified using a His tag added at its N-terminus. The enzyme demethylesterified Citrus pectin with a K m of 0.86 mg/ml. The enzyme did not require salt for activity and was found to be relatively temperature-sensitive. The precipitation of enzyme-treated pectin by CaCl2 suggested that the enzyme had a blockwise mode of pectin demethylesterification. A purified kiwi (Actinidia chinensis) pectin methylesterase inhibitor had no effect on the activity of the enzyme whereas it strongly inhibited a flax pectin methylesterase. A model of the protein structure revealed that an extra amino acid sequence in this particular Arabidopsis pectin methylesterase could form a ß-strand outside the core structure, which might be preventing the inhibitor from binding the protein.
Keywords3-D modelling Arabidopsis Cell wall Pectin methylesterase Pectin methylesterase inhibitor
Degree of esterification
Pectin methylesterase inhibitor
We thank Françoise Fournet (UPJV, Amiens) for excellent technical assistance, Prof. Catherine Sarazin (UPJV, Amiens) for helpful discussions, Dr. Matladi Ndlovu (ULB, Bruxelles) for critical reading of the manuscript, CRIHAN (centre de ressources informatiques Haute-Normandie) for molecular modelling support, and RIKEN for the AtPME31 cDNA clone.
- Ciardiello MA, D’Avino R, Amoresano A, Tuppo L, Carpentieri A, Carratore V, Tamburrini M, Giovane A, Pucci P, Camardella L (2008) The peculiar structural features of kiwi fruit pectin methylesterase: amino acid sequence, oligosaccharides structure, and modeling of the interaction with its natural proteinaceous inhibitor. Proteins 71:195–206PubMedCrossRefGoogle Scholar
- Gaffé J, Morvan C, Jauneau A, Demarty M (1992) Partial purification of flax cell wall pectin methylesterase. Phytochemistry 31:761–765Google Scholar
- Hou WC, Chang WH, Jiang CM (1999) Qualitative distinction of carboxyl group distributions in pectins with ruthenium red. Bot Bull Acad Sin 40:115–119Google Scholar
- Nicholas KB, Nicholas HB Jr, Deerfield DWII (1997) GeneDoc: analysis and visualization of genetic variation. EMBnet News 4(2):1–4Google Scholar
- Rico D, Martin-Diana AB, Barry-Ryan C, Henehan GTM, Frias JM (2007) Simultaneous modelling of the thermal degradation kinetics of pectin methylesterase in letucce (Lactuca sativa L.) and carrot (Daucus carota L.) extracts: analysis of seasonal variation and tissue type. Biosci Biotechnol Biochem 71:2383–2392PubMedCrossRefGoogle Scholar
- Willats WGT, Orfila C, Limberg G, Buchholti HC, van Alebeek GJWM, Voragen AGJ, Marcus SE, Christensen TMIE, Mikkelsen JD, Murray BS, Knox JP (2001) Modulation of the degree and pattern of methyl-esterification of pectic homogalacturonan in plant cell walls. Implications for pectin methyl esterase action, matrix properties, and cell adhesion. J Biol Chem 276:19404–19413PubMedCrossRefGoogle Scholar