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Enzyme Research
Volume 2011 (2011), Article ID 970983, 6 pages
http://dx.doi.org/10.4061/2011/970983
Research Article

Molecular Cloning and Characterization of P4 Nuclease from Leishmania infantum

1Biotechnology Research Center, Tabriz University of Medical Sciences, Tabriz 51656-65811, Iran
2Drug Applied Research Center, Tabriz University of Medical Sciences, Tabriz 51656-65811, Iran
3Infectious and Tropical Disease Research Center, Tabriz University of Medical Sciences, Tabriz 51656-65811, Iran
4Tuberculosis and Lung Disease Research Center, Tabriz University of Medical Sciences, Tabriz 51656-65811, Iran
5Deparetment of Agriculture, Zanjan University, Zanjan, Iran
6Immunology Department, Pasteur Institute of Iran, Tehran, Iran
7Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz 51656-65811, Iran

Received 3 January 2011; Revised 31 March 2011; Accepted 25 April 2011

Academic Editor: Ariel M. Silber

Copyright © 2011 Safar Farajnia et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Parasite of the genus Leishmania is reliant on the salvage pathway for recycling of ribonucleotides. A class I nuclease enzyme also known as P4 nuclease is involved in salvage of purines in cutaneous Leishmania species but the relevant enzymes have not been characterized in Leishmania infantum (L. infantum). The aim of this study was to clone and characterize the gene encoding class I nuclease in L. infantum. DNA extracted from L. infantum was used for amplification of P4 nuclease gene (Li-P4) by PCR. The product was cloned, sequenced, and expressed in E. coli for further characterization. Analysis of the sequence of Li-P4 revealed that the gene consists of an ORF of 951 bp. Sequence similarity analysis indicated that Li-P4 has a high homology to relevant enzymes of other kintoplastids with the highest homology (88%) to p1/s1 class I nuclease from L. donovani. Western blotting of antirecombinant Li-P4 with promastigote and amastigote stages of L. infantum showed that this nuclease is present in both stages of parasite with higher expression in amastigote stage. The highly conserved nature of this essential enzyme in Leishmania parasites suggests it as a promising drug target for leishmaniasis.