Plant Cytosolic Ascorbate Peroxidase with Dual Catalytic Activity Modulates Abiotic Stress Tolerances

Dan-Chu Chin; Rajendran Senthil Kumar; Ching-Shu Suen; Chia-Yu Chien; Ming-Jing Hwang; Chun-Hua Hsu; Xu Xuhan; Zhong Xiong Lai; Kai-Wun Yeh

doi:10.1016/j.isci.2019.05.014

Plant Cytosolic Ascorbate Peroxidase with Dual Catalytic Activity Modulates Abiotic Stress Tolerances

iScience. 2019 Jun 28:16:31-49. doi: 10.1016/j.isci.2019.05.014. Epub 2019 May 15.

Authors

Dan-Chu Chin¹, Rajendran Senthil Kumar¹, Ching-Shu Suen², Chia-Yu Chien³, Ming-Jing Hwang², Chun-Hua Hsu³, Xu Xuhan⁴, Zhong Xiong Lai⁴, Kai-Wun Yeh⁵

Affiliations

¹ Institute of Plant Biology, National Taiwan University, Taipei 106, Taiwan.
² Institute of Biomedical Sciences, Academia Sinica, Taipei 115, Taiwan.
³ Department of Agricultural Chemistry, National Taiwan University, Taipei 106, Taiwan.
⁴ Institute of Horticultural Biotechnology, Fujian Agriculture and Forestry University, Fuzhou, China.
⁵ Institute of Plant Biology, National Taiwan University, Taipei 106, Taiwan; Institute of Horticultural Biotechnology, Fujian Agriculture and Forestry University, Fuzhou, China. Electronic address: ykwbppp@ntu.edu.tw.

Abstract

Ascorbic acid-glutathione (AsA-GSH) cycle represents important antioxidant defense system in planta. Here we utilized Oncidium cytosolic ascorbate peroxidase (OgCytAPX) as a model to demonstrate that CytAPX of several plants possess dual catalytic activity of both AsA and GSH, compared with the monocatalytic activity of Arabidopsis APX (AtCytAPX). Structural modeling and site-directed mutagenesis identified that three amino acid residues, Pro⁶³, Asp⁷⁵, and Tyr⁹⁷, are required for oxidization of GSH in dual substrate catalytic type. Enzyme kinetic study suggested that AsA and GSH active sites are distinctly located in cytosolic APX structure. Isothermal titration calorimetric and UV-visible analysis confirmed that cytosolic APX is a heme-containing protein, which catalyzes glutathione in addition to ascorbate. Biochemical and physiological evidences of transgenic Arabidopsis overexpressing OgCytAPX1 exhibits efficient reactive oxygen species-scavenging activity, salt and heat tolerances, and early flowering, compared with Arabidopsis overexpressing AtCytAPX. Thus results on dual activity CytAPX impose significant advantage on evolutionary adaptive mechanism in planta.

Keywords: Plant Biochemistry; Plant Biology; Plant Evolution.