Triple mutation of Cys26, Trp35, and Cys126 in stromal ascorbate peroxidase confers H₂O₂ tolerance comparable to that of the cytosolic isoform

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• 作者：Sakihito Kitajima ; Masahito Kitamura ; Namiko Koja
• 关键词：Ascorbate peroxidase ; Chloroplast ; Cysteine ; Hydrogen peroxide ; Inactivation ; Radical ; Stroma ; Tryptophan
• 刊名：Biochemical and Biophysical Research Communications
• 出版年：2008
• 期刊代码：159_0006291x
• 类别：bio
• 出版时间：8 August 2008
• 卷：372
• 期：4
• 页码：918-923
• 文件大小：523 K

摘要

Ascorbate peroxidase (APX) isoforms localized in the stroma and thylakoid of the chloroplast play a principle role in detoxifying hydrogen peroxide (H₂O₂) generated in photosystem I; however, once the ascorbate is depleted, the enzyme is attacked by H₂O₂ and rapidly loses its activity. Here, we report that radical transfer across the porphyrin moiety and amino acid residues in the reaction intermediate and H₂O₂-mediated enzyme inactivation involve cooperative interactions of the Cys26, Trp35, and Cys126 residues of stromal APX. The wild-type enzyme had a half-time of inactivation of <10 s, while the triple mutant of the three residues retained 50%of the initial activity after H₂O₂ treatment for 3 min. The H₂O₂ tolerance of this mutant was comparable to that of the H₂O₂-tolerant APX isoform localized in the cytosol.