5i8u
Crystal Structure of the RV1700 (MT ADPRASE) E142Q mutant
Structural highlights
FunctionADPP_MYCTU Catalyzes the hydrolysis of ADP-ribose (ADPR) to AMP and ribose-5-phosphate. Can also hydrolyze ADP-mannose and ADP-glucose, with lower efficiency. Has weaker activity with NAD, GDP-sugars and UDP-sugars (PubMed:27683242). Also catalyzes the conversion of 8-oxo-dGDP to 8-oxo-dGMP, and 8-oxo-GDP to 8-oxo-GMP. Functions in concert with MutT1 to detoxify 8-oxo-dGTP to 8-oxo-dGMP and may play an important role in supporting cellular growth under oxidative stress (PubMed:23463507). The catalytic efficiency is much higher for the hydrolysis of ADPR than 8-oxo-dGTP, suggesting a more relevant biological role in hydrolysis of ADPR (PubMed:27683242).[1] [2] Publication Abstract from PubMedMycobacterium tuberculosis represents one of the world's most devastating infectious agents - with one third of the world's population infected and 1.5 million people dying each year from this deadly pathogen. As part of an effort to identify targets for therapeutic intervention, we carried out the kinetic characterization of the product of gene rv1700 of M. tuberculosis. Based on its sequence and its structure, the protein had been tentatively identified as a pyrophosphohydrolase specific for adenosine diphosphate ribose (ADPR), a compound involved in various pathways including oxidative stress response and tellurite resistance. In this work we carry out a kinetic, mutational and structural investigation of the enzyme, which provides a full characterization of this Mt-ADPRase. Optimal catalytic rates were achieved at alkaline pH (7.5-8.5) with either 0.5-1 mM Mg2+ or 0.02-1 mM Mn2+. K m and k cat values for hydrolysis of ADPR with Mg2+ ions are 200 +/- 19 muM and 14.4 +/- 0.4 s-1, and with Mn2+ ions are 554 +/- 64 muM and 28.9 +/- 1.4 s-1. Four residues proposed to be important in the catalytic mechanism of the enzyme were individually mutated and the kinetics of the mutant enzymes were characterized. In the four cases, the K m increased only slightly (2- to 3-fold) but the k cat decreased significantly (300- to 1900-fold), confirming the participation of these residues in catalysis. An analysis of the sequence and structure conservation patterns in Nudix ADPRases permits an unambiguous identification of members of the family and provides insight into residues involved in catalysis and their participation in substrate recognition in the Mt-ADPRase. Kinetic and mutational studies of the adenosine diphosphate ribose hydrolase from Mycobacterium tuberculosis.,O'Handley SF, Thirawatananond P, Kang LW, Cunningham JE, Leyva JA, Amzel LM, Gabelli SB J Bioenerg Biomembr. 2016 Sep 28. PMID:27683242[3] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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