**6. References**

288 Understanding Tuberculosis – Deciphering the Secret Life of the Bacilli

one of the cleft. Although crystal structure of PZA-bound PncA is not known but it is suggested that the binding of PZA involves hydrogen bonded interaction between pyridyl nitrogen atom of PZA and the conserved water molecule W1 which is coordinated to Fe2+ ion. Another possible interaction is provided by carbonyl oxygen atom of PZA with peptide Ala134 – Cys138. Overall, the role played by Fe2+ ion and His57 is critical in stabilizing the structure of substrate-binding site. Finally the conversion of PZA to POA and its accumulation results in lowering the intracellular pH to suboptimal level and thus causing the inactivation of some of the critically important proteins such as fatty acid synthase

Both prodrugs INH and PZA bind to LPO in the substrate-binding site on the distal heme side. In the unliganded structure of LPO, the space of the substrate-binding site is filled with six water molecules, W1, W2, W3, W4¸ W5 and W6. Upon binding to INH, four water molecules W2, W4¸ W5 and W6 are expelled from the site. In the structure of the complex of LPO with PZA, three water molecules W4, W5 and W6 are displaced. The conserved water molecule W1 occupies a position on distal heme side in the centre between the positions of heme-iron and Nδ2 atom of His109. His109 is linked to a chain of six other conserved water molecules, W1, W2, W3, W4¸ W5 and W6 together with His266 and Asp253 residues interlinked between them. The pyridine ring nitrogen atom is hydrogen bonded to W1 which in turn is hydrogen bonded to Nδ2 of His109 and heme iron. The position and orientation of INH are fixed in the substrate-binding site because of a hydrogen bonded interaction between amino nitrogen atom of INH and N2 of Gln423. The position of PZA also fixed in the substrate-binding site on the distal heme side where amino nitrogen atom of PZA forms a hydrogen bond with water molecule W2 which in turn is hydrogen bonded to the conserved water molecule W1. As in the complex of LPO with INH, W1 is hydrogen bonded to heme iron and His109 Nδ2. On the other side carbonyl oxygen atom of PZA is hydrogen bonded to Gln423 N2. However, unlike INH where pyridine nitrogen atom is hydrogen bonded to W1, in this case carboxamide nitrogen atom is hydrogen bonded to W1 via another water molecule W2. The main difference between the two complexes is that the INH molecule interacts directly with heme water molecule W1 while PZA binds to heme

As far as the ligand binding sites are concerned all the three enzymes, LPO, *Mt*CP and PncA show considerable similarities with strong preferences for the binding of small aromatic compounds. There is a clear evidence that INH is converted in a similar manner into active form by both LPO and *Mt*CP indicating a direct role of LPO in the treatment of TB. Similarly, PZA also makes a good substrate for LPO which can be converted into active form as an antibacterial agent. However, the final active forms produced by LPO and PncA may not be same and the mechanism of action may be different. Nevertheless, LPO seems to have a role in the treatment of tuberculosis through its interactions with INH and PZA and

The authors acknowledge Department of Science and Technology (DST) of the Central Government, New Delhi for the financial support. NP and MS thank the Indian Council of

resulting in the killing of bacteria (Zimhony et al., 2000).

water molecule W1 via another water molecule W2.

it should be exploited.

**5. Acknowledgements** 

**4. Discussion** 


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**16** 

*Uruguay* 

**Thiol-Dependent Peroxidases in** 

Martín Hugo1,2, Rafael Radi1,2 and Madia Trujillo1,2

*Mycobacterium tuberculosis* (*M. tuberculosis*) is the causative agent of tuberculosis disease. According to the Global Tuberculosis Control 2010 report of the World Health Organization http://www.who.int/tb/publications/global\_report/en/, approximately one-third of the world's population is latently infected with *M. tuberculosis* and about two million people die of this disease every year. The emergence of multi- and extensively- drug resistant strains to the currently available drugs makes the development of new therapeutic strategies a priority. However, the mechanisms underlying pathogenesis, virulence and persistence of infections caused by *M. tuberculosis* are not completely understood (Nathan, 2009; Lawn &

*M. tuberculosis* is one of the most successful human pathogens. It has evolved diverse strategies to ensure growth and survival inside the hostile environment of macrophages, its primary host cells (Ehrt & Schnappinger, 2009; Meena & Rajni, 2010). The molecular mechanisms of *M. tuberculosis* pathogenesis are under active investigation, since they could provide the basis for a rationalized drug design. These include inhibition of phagosome maturation into phagolysosomes (Armstrong & Hart, 1971; MacMicking *et al.*, 2003), inhibition of the acidification of *Mycobacterium-*harboring phagosomes (Sturgill-Koszycki *et al.*, 1994), DNA repair and protein repair or degradation (Boshoff *et al.*, 2003; Gandotra *et al.*, 2007; Lee *et al.*, 2009), as well as decomposition of cytotoxic reactive nitrogen and oxygen species formed upon phagocytosis (Nathan & Shiloh, 2000; Shiloh & Nathan, 2000; Bedard & Krause, 2007). These should be considered as complementary survival mechanisms. Herein, we will focus in the antioxidant systems of *M. tuberculosis*, and particularly, in thiol-

**2. Formation of reactive oxygen and nitrogen species by activated** 

Upon phagocytosis, NADPH oxidase (NADPHox) assembles into an enzymatically active complex that transfers electrons from NADPH to molecular oxygen producing superoxide

**1. Introduction** 

Zumla, 2011).

dependent peroxidases.

**macrophages** 

*Mycobacterium tuberculosis*

*2Center for Free Radical and Biomedical Research, Facultad de Medicina, Universidad de la República,* 

**Antioxidant Defense** 

*1Departamento de Bioquímica* 

