Sandbox Reserved 1068: Difference between revisions
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==Introduction== | ==Introduction== | ||
Salicylate synthase from ''Mycobacterim tuberculosis'' (MtbI) is a highly promiscuous enzyme that has four distinct activities ''in vivo'': isochorismate synthase (IS), isochorismate pyruvate lyase (IPL), salicylate synthase (SS) and chromate mutate (CM). MtbI belongs to the chorismate-utilising enzyme family, which consists of structural homologues (<scene name='69/694235/Irp9/2'>Ipr9</scene>, <scene name='69/694235/Menf/2'>MenF</scene>, <scene name='69/694235/Entc/2'>EntC</scene>, and <scene name='69/694235/Mbti/2'>MbtI</scene>) that isomerize chromate to isochorismate. These enzymes are present in bacteria, fungi, plants and apicomplexan parasites and catalyze the initial reactions of menaquinone, siderophore, and tryptophan biosynthesis. The IS, IPL, and SS activity of MbtI require the presence of a magnesium ion within the active site, while CM activity is only observed in absence of the magnesium cation. IS, IPL, and SS activity are also modulated by the pH of the medium. Isochorismate is the primary product at pH values below 7.5 and salicylate is the primary product formed at pH 8. | Salicylate synthase from ''Mycobacterim tuberculosis'' (MtbI) is a highly promiscuous enzyme that has four distinct activities ''in vivo'': isochorismate synthase (IS), isochorismate pyruvate lyase (IPL), salicylate synthase (SS) and chromate mutate (CM). MtbI belongs to the chorismate-utilising enzyme family, which consists of structural homologues (<scene name='69/694235/Irp9/2'>Ipr9</scene>, <scene name='69/694235/Menf/2'>MenF</scene>, <scene name='69/694235/Entc/2'>EntC</scene>, and <scene name='69/694235/Mbti/2'>MbtI</scene>) that isomerize chromate to isochorismate. These enzymes are present in bacteria, fungi, plants and apicomplexan parasites and catalyze the initial reactions of menaquinone, siderophore, and tryptophan biosynthesis. The IS, IPL, and SS activity of MbtI require the presence of a magnesium ion within the active site, while CM activity is only observed in absence of the magnesium cation. IS, IPL, and SS activity are also modulated by the pH of the medium. Isochorismate is the primary product at pH values below 7.5 and salicylate is the primary product formed at pH 8. | ||
The salicylate synthase activity of MbtI catalyzes the first committed step in the synthesis of the iron chelating siderophore, mycobactin, in ''Mycobacterium tuberculosis'' (Figure 3)<ref name= " | The salicylate synthase activity of MbtI catalyzes the first committed step in the synthesis of the iron chelating siderophore, mycobactin, in ''Mycobacterium tuberculosis'' (Figure 3)<ref name= "5a">PMID:22607697</ref>. This complex secondary metabolite is essential for both virulence and survival of ''M. tuberculosis''. Therefore, inhibitors of salicylate synthase may serve as potential TB therapies with a novel mode of action <ref name= "1a"> DOI: 10.1002/cmdc/201000137</ref> <ref name= "2a">PMID:23108268</ref> <ref name= "7a">Voss, James J., Kerry Rutter, Benjamin G. Schroedor, Hua Su, and YaQi Zhu. "The salicylate-derived mycobactin siderophores of Mycobacterium tuberculosis are essential for growth in macrophages." Proceedings of the National Academy of Sciences 97.3 (2000): 1252-57. Web. 14 Mar. 2015.</ref> <ref name= "5a"/> <ref name= "4a">DOI:10.1021/bi2009739</ref> <ref name= "9a">PMID:14982443</ref> (1,2,7,5,4,9) | ||
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''Mycobacterium tuberculosis'' is the causative agent of Tuberculosis (TB), an infectious disease that affects one-third of the worlds population. Two TB-related conditions exist: latent TB infection and active TB disease. Currently, there are four regimens that are approved for the treatment of latent TB infection through the use of the antibiotics isoniazid, rifampin, and rifapentine.TB disease can also be treated through various antibiotic regimens. There are 10 drugs currently approved by the FDA for treating TB disease. The first-line anti-TB agents are the antibiotics isoniazid, rifampin, ethambutol, and pyrazinamide <ref>Tuberculosis (TB). Ed. Sam Posner. Centers for Disease Control and Prevention, n.d. Web. 9 Apr. 2015.</ref>. Although various treatments for TB infection and TB disease exist, the emergence of multi-drug and extensively-drug resistant strains of ''M. tuberculosis'' has increased the need for anti-tubercular agents with novel modes of action. | ''Mycobacterium tuberculosis'' is the causative agent of Tuberculosis (TB), an infectious disease that affects one-third of the worlds population. Two TB-related conditions exist: latent TB infection and active TB disease. Currently, there are four regimens that are approved for the treatment of latent TB infection through the use of the antibiotics isoniazid, rifampin, and rifapentine.TB disease can also be treated through various antibiotic regimens. There are 10 drugs currently approved by the FDA for treating TB disease. The first-line anti-TB agents are the antibiotics isoniazid, rifampin, ethambutol, and pyrazinamide <ref>Tuberculosis (TB). Ed. Sam Posner. Centers for Disease Control and Prevention, n.d. Web. 9 Apr. 2015.</ref>. Although various treatments for TB infection and TB disease exist, the emergence of multi-drug and extensively-drug resistant strains of ''M. tuberculosis'' has increased the need for anti-tubercular agents with novel modes of action. | ||
Iron is essential for mycobacterial growth and pathogenesis, therefore the pathways for iron acquisition are potential targets for antibacterial therapies.''M. tuberculosis'' obtains iron through two different pathways: chelating iron from the host through the siderophore mycobactin and the degradation of heme released from damaged red blood cells'''.'''Mycobactin is a siderophore synthesized by the proteins encoded by the ''mbt'' and ''mbt2'' gene cluters <ref name=" | Iron is essential for mycobacterial growth and pathogenesis, therefore the pathways for iron acquisition are potential targets for antibacterial therapies.''M. tuberculosis'' obtains iron through two different pathways: chelating iron from the host through the siderophore mycobactin and the degradation of heme released from damaged red blood cells'''.'''Mycobactin is a siderophore synthesized by the proteins encoded by the ''mbt'' and ''mbt2'' gene cluters <ref name="5a"/>. MbtI is the first enzyme in the mycobactin biosynthesis pathway and is a potential target for inhibition. The salicylate synthase activity of MbtI produces salicylate and pyruvate from chorismate through an isochorismate intermediate. Inhibition of MbtI activity would decrease the production of salicylate and therefore the synthesis of mycobactin; leading to a decrease in iron acquisition and pathogenesis of ''M. tuberculosis''<ref>De Voss, James J., Kerry Rutter, Benjamin G. Schroeder, Hua Su, and YaQi Zhu. The salicylate-derived mycobacterium siderophore of Mycobacterium tuberculosis are essential for growth in macrophages. "Proceedings of the National Science Academy" 97.3 (2000): 1252-57. Web. 5 Apr. 2015.</ref> . | ||
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== Structural highlights == | == Structural highlights == | ||
MtI structure has a mobile element (residues 323 to 227) that can adopt a <scene name='69/694235/Irp9_closed_state/2'>closed</scene> or <scene name='69/694235/2g5f_with_open_loop/1'>open conformation</scene> depending on whether or not ligands are bound to the active site. The closed conformation partially obstructs the active site. <ref name= " | MtI structure has a mobile element (residues 323 to 227) that can adopt a <scene name='69/694235/Irp9_closed_state/2'>closed</scene> or <scene name='69/694235/2g5f_with_open_loop/1'>open conformation</scene> depending on whether or not ligands are bound to the active site. The closed conformation partially obstructs the active site. <ref name= "5a"/>. | ||
Inhibition studies have also shown a switch in binding mode at the MbtI active site for inhibitors carrying a substituted enolpyruvyl group compared to the chorismate substrate. Crystal structures and fluorescent-based thermal shift assays show that substituents larger than a methyl group are accommodated in the active site of MbtI through localized flexibility in the peptide backbone. Positioning of the active site residues of MbtI with the inhibitor AMT is highly similar to the closed form of MbtI <ref name= " | Inhibition studies have also shown a switch in binding mode at the MbtI active site for inhibitors carrying a substituted enolpyruvyl group compared to the chorismate substrate. Crystal structures and fluorescent-based thermal shift assays show that substituents larger than a methyl group are accommodated in the active site of MbtI through localized flexibility in the peptide backbone. Positioning of the active site residues of MbtI with the inhibitor AMT is highly similar to the closed form of MbtI <ref name= "5a"/>. | ||
==Molecular Mechanism== | ==Molecular Mechanism== | ||
