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Soybean seed pectinesterase

Published online by Cambridge University Press:  19 September 2008

Oskar Markovič and
Ralph L. Obendorf
Oskar Markovič
Affiliation:
Seed Biology, Department of Soil, Crop and Atmospheric Sciences, Cornell University Agricultural Experiment Station, 619 Bradfield Hall, Cornell University, Ithaca, NY 14853-1901, USAInstitute of Chemistry, Slovak Academy of Sciences, Dúbravská cesta 9, SK 842 38 Bratislava, Slovakia
Ralph L. Obendorf*
Affiliation:
Seed Biology, Department of Soil, Crop and Atmospheric Sciences, Cornell University Agricultural Experiment Station, 619 Bradfield Hall, Cornell University, Ithaca, NY 14853-1901, USA
*
*Fax: +1 607 255 2644 E-mail: rlol@cornell.edu
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Abstract

Methanol accumulates in axis tissues of maturing soybean seeds, correlating with preharvest seed deterioration. Accumulation of methanol appears to be associated with the enzymic demethylation of pectin methyl esters by pectinesterase (PE; EC 3.1.1.11). To characterize PE in developing and maturing soybean (Glycine max (L.) Merrill) seeds, enzyme activity was assayed in axis and cotyledon tissues. Activity per g fresh weight was 20–25 times higher in axes than in cotyledons with highest activities between 45 and 60 days after flowering (DAF). Twenty to 33% of the total PE activity was in the ‘soluble’ form (extracted with water, 0.5 M sucrose, 1 M sucrose and water). Soluble and cell-wall-bound PE (subsequently extracted with 1 M NaCI) were purified and characterized from axes of seeds at 45–60 DAF. Purification of PE was achieved through concentration of extracts by ultra-filtration, precipitation with ammonium sulfate (30–80% saturation), dialysis, gel filtration on Sephadex G-75 columns, and ion exchange chromatography on CM Sepharose CL-6B. Further purification of both soluble and bound PE was by isoelectric focusing (IEF) on ultrathin layers of polyacrylamide gel with simultaneous detection of protein and PE activity. It was possible to follow seven bands exhibiting PE activity with pl values between 6.0 and 9.5 in 1 M NaCI-extracts of total homogenates. Differences in the IEF patterns of bound and soluble PE were observed. Whereas the bound enzyme exhibited more basic PE bands (pl 8–9.5), the soluble enzyme had more active bands at pl 6.5, 7.0 and 7.5. The Mr was close to 33 000 and the pH optimum was 7.8 for both soluble and bound PE.

Keywords

cell wallenzyme activityisoelectric focusing (IEF)pectinesterase (EC 3.1.1.11)seed developmentsoybean (Glycine max (L.) Merr.)

Information

Type
Physiology & Biochemistry
Information
Seed Science Research , Volume 8 , Issue 4 , December 1998, pp. 455 - 461
Copyright
Copyright © Cambridge University Press 1998

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