Modulation of the Activity of Mycobacterium tuberculosis LipY by Its PE Domain.

Christopher K Garrett,Saskia B Neher,Lindsey J Broadwell,Cassandra K Hayne,Giovanni Delogu

doi:10.1371/journal.pone.0135447

Christopher K Garrett, Saskia B Neher + Show 3 more

Open Access

https://doi.org/10.1371/journal.pone.0135447

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Journal: PloS one	Publication Date: Aug 13, 2015
Citations: 21	License type: CC BY 4.0

Affiliation: University of North Carolina at Chapel Hill

Abstract

Mycobacterium tuberculosis harbors over 160 genes encoding PE/PPE proteins, several of which have roles in the pathogen’s virulence. A number of PE/PPE proteins are secreted via Type VII secretion systems known as the ESX secretion systems. One PE protein, LipY, has a triglyceride lipase domain in addition to its PE domain. LipY can regulate intracellular triglyceride levels and is also exported to the cell wall by one of the ESX family members, ESX-5. Upon export, LipY’s PE domain is removed by proteolytic cleavage. Studies using cells and crude extracts suggest that LipY’s PE domain not only directs its secretion by ESX-5, but also functions to inhibit its enzymatic activity. Here, we attempt to further elucidate the role of LipY’s PE domain in the regulation of its enzymatic activity. First, we established an improved purification method for several LipY variants using detergent micelles. We then used enzymatic assays to confirm that the PE domain down-regulates LipY activity. The PE domain must be attached to LipY in order to effectively inhibit it. Finally, we determined that full length LipY and the mature lipase lacking the PE domain (LipYΔPE) have similar melting temperatures. Based on our improved purification strategy and activity-based approach, we concluded that LipY’s PE domain down-regulates its enzymatic activity but does not impact the thermal stability of the enzyme.

Highlights

Mycobacterium tuberculosis (Mtb) is remarkably adept at interfering with host cellular processes in order to evade destruction
PE/PPE proteins are unique to mycobacteria; they were initially discovered when sequencing of the Mtb genome revealed approximately 160 genes encoding proteins with Pro–Glu (PE) or Pro–Pro–Glu (PPE) motifs near their N-termini[4]
Studies with individual PE/PPE proteins suggest that they serve to target their C-terminal cargo to the type VII secretion systems[19, 36, 37]

Summary

Introduction

Mycobacterium tuberculosis (Mtb) is remarkably adept at interfering with host cellular processes in order to evade destruction. This ability depends on the secretion of virulence factors that modify the host environment. Known as PE and PPE proteins, are involved in immune evasion and virulence[1,2,3]. PE/PPE proteins are unique to mycobacteria; they were initially discovered when sequencing of the Mtb genome revealed approximately 160 genes encoding proteins with Pro–Glu (PE) or Pro–Pro–Glu (PPE) motifs near their N-termini[4]. Subsequent analysis revealed that PE/PPE proteins comprise about 7% of the coding capacity of the Mtb genome[5]. PE/PPE domains have been identified in both pathogenic and saprophytic mycobacteria, pathogenic mycobacteria maintain the highest number of PE/PPE proteins[6]

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