Photosynthetica 2000, 38(1):45-52 | DOI: 10.1023/A:1026739806712

Purification and Stability during Storage of Phoshoenolpyruvate Carboxylase from Leaves of Amaranthus hypochondriacus, a NAD-ME Type C4 Plant

J. Gayathri1, K. Parvathi1, A.S. Raghavendra2
1 Department of Plant Sciences, School of Life Sciences, University of Hyderabad, Hyderabad, India
2 Department of Plant Sciences, School of Life Sciences, University of Hyderabad, Hyderabad, India

A traditional method is reported for purification of phosphoenolpyruvate carboxylase (PEPC; EC 4.1.1.31) from leaves of Amaranthus hypochondriacus L. with a high yield of 50 %, 135-fold purification, and specific activity of 900 mmol kg-1(protein) s-1. PEPC was purified from light-adapted leaves of A. hypochondriacus, involving 40-60 % ammonium sulphate fractionation, followed by chromatography on columns of DEAE-Sepharose, hydroxylapatite (HAP), and Seralose 6-B. The enzyme appeared as a single band on 10 % SDS-PAGE, with a molecular mass of about 100 kDa. Kinetic studies with purified enzyme confirmed the PEPC to be the light-form of the enzyme. Glycerol generally increased the stability of PEPC. The stability and storage of the purified enzyme was studied at temperatures of 4 °C, -20 °C, and liquid nitrogen. PEPC maintained its activity for up to 3 months upon storage with 50 % (v/v) glycerol in liquid nitrogen.

Additional key words: DEAE-Sepharose; glucose-6-phosphate; glycerol; hydroxylapatite; L-malate; phosphate; phosphoenolpyruvate; Seralose- 6B

Prepublished online: October 1, 2000; Published: April 1, 2000  Show citation

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Gayathri, J., Parvathi, K., & Raghavendra, A.S. (2000). Purification and Stability during Storage of Phoshoenolpyruvate Carboxylase from Leaves of Amaranthus hypochondriacus, a NAD-ME Type C4 Plant. Photosynthetica, 38(1), 45-52. doi: 10.1023/A:1026739806712
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