Mycobacterium tuberculosis Prokaryotic Ubiquitin-like Protein-deconjugating Enzyme Is an Unusual Aspartate Amidase

Deamidase of Pup (Dop), the prokaryotic ubiquitin-like protein (Pup)-deconjugating enzyme, is critical for the full virulence of Mycobacterium tuberculosis and is unique to bacteria, providing an ideal target for the development of selective chemotherapies. We used a combination of genetics and chem...

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Bibliographic Details
Published in:The Journal of biological chemistry 2012-10, Vol.287 (44), p.37522-37529
Main Authors: Burns, Kristin E., McAllister, Fiona E., Schwerdtfeger, Carsten, Mintseris, Julian, Cerda-Maira, Francisca, Noens, Elke E., Wilmanns, Matthias, Hubbard, Stevan R., Melandri, Francesco, Ovaa, Huib, Gygi, Steven P., Darwin, K.Heran
Format: Article
Language:English
Subjects:
Amidohydrolases - chemistry
Amidohydrolases - genetics
Amino Acid Motifs
Amino Acid Sequence
Amino Acid Substitution
Bacterial Proteins - chemistry
Bacterial Proteins - genetics
Biocatalysis
Catalytic Domain
Depupylation
Diazooxonorleucine - chemistry
Enzyme Inhibitors - chemistry
Hydrolysis
Models, Molecular
Molecular Sequence Data
Mutagenesis, Site-Directed
Mycobacterium tuberculosis
Mycobacterium tuberculosis - enzymology
Protease
Proteasome
Protein Deamidation
Protein Degradation
Protein Synthesis and Degradation
Pup
Structural Homology, Protein
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Summary:Deamidase of Pup (Dop), the prokaryotic ubiquitin-like protein (Pup)-deconjugating enzyme, is critical for the full virulence of Mycobacterium tuberculosis and is unique to bacteria, providing an ideal target for the development of selective chemotherapies. We used a combination of genetics and chemical biology to characterize the mechanism of depupylation. We identified an aspartate as a potential nucleophile in the active site of Dop, suggesting a novel protease activity to target for inhibitor development. Background: Dop is critical for the full virulence of Mycobacterium tuberculosis; however, its mechanism is not understood. Results: Asp-95 was identified as a catalytically significant residue. Conclusion: This work suggests that Asp-95 functions either as a direct nucleophile forming a unique anhydride intermediate or is part of a catalytic center that includes polarized water as the nucleophile. Significance: Understanding the mechanism of Dop can help guide the design and selection of inhibitors.
ISSN:0021-9258
1083-351X
DOI:10.1074/jbc.M112.384784