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Characterization and inhibition of a class II diterpene cyclase from Mycobacterium tuberculosis: implications for tuberculosis.

Mann FM, Prisic S, Hu H, Xu M, Coates RM, Peters RJ - J. Biol. Chem. (2009)

Bottom Line: Chem.Soc., in press).Although arguably not suitable for clinical use, these nevertheless provide a basis for pharmaceutical design against this intriguing biosynthetic pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Iowa State University, Ames, Iowa 50011, USA.

ABSTRACT
Mycobacterium tuberculosis remains a widespread and devastating human pathogen, whose ability to infiltrate macrophage host cells from the human immune system is an active area of investigation. We have recently reported the discovery of a novel diterpene from M. tuberculosis, edaxadiene, whose ability to arrest phagosomal maturation in isolation presumably contributes to this critical process in M. tuberculosis infections. (Mann, F. M., Xu, M., Chen, X., Fulton, D. B., Russell, D. G., and Peters, R. J. (2009) J. Am. Chem. Soc., in press). Here, we present characterization of the class II diterpene cyclase that catalyzes the committed step in edaxadiene biosynthesis, i.e. the previously identified halimadienyl-diphosphate synthase (HPS; EC 5.5.1.16). Intriguingly, our kinetic analysis suggests a potential biochemical regulatory mechanism that triggers edaxadiene production upon phagosomal engulfment. Furthermore, we report characterization of potential HPS inhibitors: specifically, two related transition state analogs (15-aza-14,15-dihydrogeranylgeranyl diphosphate (7a) and 15-aza-14,15-dihydrogeranylgeranyl thiolodiphosphate (7b)) that exhibit very tight binding. Although arguably not suitable for clinical use, these nevertheless provide a basis for pharmaceutical design against this intriguing biosynthetic pathway. Finally, we provide evidence indicating that this pathway exists only in M. tuberculosis and is not functional in the closely related Mycobacterium bovis because of an inactivating frameshift in the HPS-encoding gene. Thus, we hypothesize that the inability to produce edaxadiene may be a contributing factor in the decreased infectivity and/or virulence of M. bovis relative to M. tuberculosis in humans.

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Outline of chemical syntheses of aza analog inhibitors 15-aza-GGPP (7a) and 15-aza-GGSPP (7b) from 14,15-epoxy-GGOH (5) via a common 15-aza-GGOH intermediate (6a).
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Figure 2: Outline of chemical syntheses of aza analog inhibitors 15-aza-GGPP (7a) and 15-aza-GGSPP (7b) from 14,15-epoxy-GGOH (5) via a common 15-aza-GGOH intermediate (6a).

Mentions: Initial functional characterization of HPS was limited by enzymatic instability. Here, we report the development of a construct amendable to kinetic characterization, along with the implications of the observed striking Mg2+ cofactor inhibition effect. We further report analysis of potential inhibitors, with two transition state analogs (7a and 7b) (Fig. 2) found to exhibit high affinity. In addition, investigation of the corresponding gene in M. bovis demonstrates the presence of an inactivating frameshift, abrogating the ability of this otherwise closely related mycobacterium to produce edaxadiene (4), which we hypothesize contributes to its reduced infectivity and/or virulence in humans relative to M. tuberculosis.


Characterization and inhibition of a class II diterpene cyclase from Mycobacterium tuberculosis: implications for tuberculosis.

Mann FM, Prisic S, Hu H, Xu M, Coates RM, Peters RJ - J. Biol. Chem. (2009)

Outline of chemical syntheses of aza analog inhibitors 15-aza-GGPP (7a) and 15-aza-GGSPP (7b) from 14,15-epoxy-GGOH (5) via a common 15-aza-GGOH intermediate (6a).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2749132&req=5

Figure 2: Outline of chemical syntheses of aza analog inhibitors 15-aza-GGPP (7a) and 15-aza-GGSPP (7b) from 14,15-epoxy-GGOH (5) via a common 15-aza-GGOH intermediate (6a).
Mentions: Initial functional characterization of HPS was limited by enzymatic instability. Here, we report the development of a construct amendable to kinetic characterization, along with the implications of the observed striking Mg2+ cofactor inhibition effect. We further report analysis of potential inhibitors, with two transition state analogs (7a and 7b) (Fig. 2) found to exhibit high affinity. In addition, investigation of the corresponding gene in M. bovis demonstrates the presence of an inactivating frameshift, abrogating the ability of this otherwise closely related mycobacterium to produce edaxadiene (4), which we hypothesize contributes to its reduced infectivity and/or virulence in humans relative to M. tuberculosis.

Bottom Line: Chem.Soc., in press).Although arguably not suitable for clinical use, these nevertheless provide a basis for pharmaceutical design against this intriguing biosynthetic pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Iowa State University, Ames, Iowa 50011, USA.

ABSTRACT
Mycobacterium tuberculosis remains a widespread and devastating human pathogen, whose ability to infiltrate macrophage host cells from the human immune system is an active area of investigation. We have recently reported the discovery of a novel diterpene from M. tuberculosis, edaxadiene, whose ability to arrest phagosomal maturation in isolation presumably contributes to this critical process in M. tuberculosis infections. (Mann, F. M., Xu, M., Chen, X., Fulton, D. B., Russell, D. G., and Peters, R. J. (2009) J. Am. Chem. Soc., in press). Here, we present characterization of the class II diterpene cyclase that catalyzes the committed step in edaxadiene biosynthesis, i.e. the previously identified halimadienyl-diphosphate synthase (HPS; EC 5.5.1.16). Intriguingly, our kinetic analysis suggests a potential biochemical regulatory mechanism that triggers edaxadiene production upon phagosomal engulfment. Furthermore, we report characterization of potential HPS inhibitors: specifically, two related transition state analogs (15-aza-14,15-dihydrogeranylgeranyl diphosphate (7a) and 15-aza-14,15-dihydrogeranylgeranyl thiolodiphosphate (7b)) that exhibit very tight binding. Although arguably not suitable for clinical use, these nevertheless provide a basis for pharmaceutical design against this intriguing biosynthetic pathway. Finally, we provide evidence indicating that this pathway exists only in M. tuberculosis and is not functional in the closely related Mycobacterium bovis because of an inactivating frameshift in the HPS-encoding gene. Thus, we hypothesize that the inability to produce edaxadiene may be a contributing factor in the decreased infectivity and/or virulence of M. bovis relative to M. tuberculosis in humans.

Show MeSH
Related in: MedlinePlus