Leishmania major encodes an unusual peroxidase that is a close homolog of plant ascorbate peroxidase: a novel role of the transmembrane domain

Adak, Subrata ; Datta, Alok K. (2005) Leishmania major encodes an unusual peroxidase that is a close homolog of plant ascorbate peroxidase: a novel role of the transmembrane domain Biochemical Journal, 390 . pp. 465-474. ISSN 0264-6021

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Official URL: http://www.biochemj.org/bj/390/bj3900465.htm

Related URL: http://dx.doi.org/10.1042/BJ20050311

Abstract

Haem containing enzymes (peroxidase and catalase) are widely distributed among prokaryotes and eukaryotes and play a vital role in H2O2-detoxification. But to date no haem containing enzymatic defense against toxic-H2O2 has been discovered in Leishmania species. We cloned, expressed and purified an unusual plant like ascorbate peroxidase from L. major (LmAPX) and characterized its catalytic parameters under steady state condition. Examination of its protein sequence indicated approx. 30- 60% identity to other ascorbate peroxidases. The N-terminus extension of LmAPX is characterized by a charged region followed by a stretch of 22 amino acid containing a transmembrane domain. To understand how the transmembrane domain influences the structure-function of LmAPX, we generated, purified and extensively characterized a variant that lacked the transmembrane domain. Eliminating the transmembrane domain had no impact on substrate binding affinity but slowed ascorbate oxidation and increased resistance to H2O2 dependent inactivation in absence of electron donor by 480-folds. Spectral studies show that H2O2 can quickly oxidize native enzyme to compound II, which subsequently is reduced back to the native enzyme by electron donor. In contrast, ascorbate free-transmembrane domain containing enzyme did not react with H2O2, as revealed by absence of compound II formation. Our findings suggest that the single copy LmAPX gene may play an important role in detoxification of H2O2 that is generated by endogenous processes and as a result of external influences such as oxidative burst condition of infected host macrophage or during drug metabolism by Leishmania.

Item Type:Article
Source:Copyright of this article belongs to Portland Press Limited.
Keywords:Ascorbate Peroxidase; Guaiacol; Hydrogen Peroxide (HO); Leishmania Major; Promastigote; Transmembrane Domain
ID Code:9083
Deposited On:29 Oct 2010 11:42
Last Modified:16 May 2016 18:56

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