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Mycobacterium tuberculosis can reside and persist in deep tissues; latent tuberculosis can evade immune detection and has a unique mechanism to convert it into active disease through reactivation. M. tuberculosis Rv1421 (MtRv1421) is a hypothetical protein that has been proposed to be involved in nucleotide binding-related metabolism in cell-growth and cell-division processes. However, due to a lack of structural information, the detailed function of MtRv1421 remains unclear. In this study, a truncated N-terminal domain (NTD) of MtRv1421, which contains a Walker A/B-like motif, was purified and crystallized using PEG 400 as a precipitant. The crystal of MtRv1421-NTD diffracted to a resolution of 1.7 Å and was considered to belong to either the C-centered monoclinic space group C2 or the I-centered orthorhombic space group I222, with unit-cell parameters a = 124.01, b = 58.55, c = 84.87 Å, β = 133.12° or a = 58.53, b = 84.86, c = 90.52 Å, respectively. The asymmetric units of the C2 or I222 crystals contained two or one monomers, respectively. In terms of the binding ability of MtRv1421-NTD to various ligands, uridine diphosphate (UDP) and UDP-N-acetylglucosamine significantly increased the melting temperature of MtRv1421-NTD, which indicates structural stabilization through the binding of these ligands. Altogether, the results reveal that a UDP moiety may be required for the interaction of MtRv1421-NTD as a nucleotide-binding protein with its ligand., (open access.) more...

Mycobacterium tuberculosis possesses the ability to undergo physiological adaptations in order to persist during the prolonged course of infection despite the active immune response of the host and in order to overcome multiple environmental changes. Previous studies have proposed that M. tuberculosis CuvA (Rv1422; MtCuvA) might play a critical role in the adaptation of the bacterium to environmental changes, such as nutrient utilization and alteration of the growth rate. However, the detailed function of MtCuvA still remains unclear owing to a lack of structural information. To better understand its role in host adaptation, MtCuvA was purified to homogeneity and was crystallized for the first time using the hanging-drop vapor-diffusion method. The crystal of MtCuvA diffracted to a resolution of 2.1 Å and belonged to the orthorhombic space group P2 1 2 1 2 1 , with unit-cell parameters a = 47.27, b = 170.93, c = 178.10 Å. The calculated Matthews coefficient (V M ) was 2.4 Å 3  Da -1 , with a solvent content of 48.02%, and thus four molecules appeared to be present in the asymmetric unit. Moreover, it is reported that MtCuvA can bind to the cell-wall precursor components uridine diphosphate (UDP)-glucose and UDP-N-acetylglucosamine. more...

GroEL is a group I chaperonin that facilitates protein folding and prevents protein aggregation in the bacterial cytosol. Mycobacteria are unusual in encoding two or more copies of GroEL in their genome. While GroEL2 is essential for viability and likely functions as the general housekeeping chaperonin, GroEL1 is dispensable, but its structure and function remain unclear. Here, we present the 2.2-Šresolution crystal structure of a 23-kDa fragment of Mycobacterium tuberculosis GroEL1 consisting of an extended apical domain. Our X-ray structure of the GroEL1 apical domain closely resembles those of Escherichia coli GroEL and M. tuberculosis GroEL2, thus highlighting the remarkable structural conservation of bacterial chaperonins. Notably, in our structure, the proposed substrate-binding site of GroEL1 interacts with the N-terminal region of a symmetry-related neighboring GroEL1 molecule. The latter is consistent with the known GroEL apical domain function in substrate binding and is supported by results obtained from using peptide array technology. Taken together, these data show that the apical domains of M. tuberculosis GroEL paralogs are conserved in three-dimensional structure, suggesting that GroEL1, like GroEL2, is a chaperonin., (Copyright © 2010 Elsevier Ltd. All rights reserved.) more...

Full-length GroEL1 from Mycobacterium tuberculosis H37Rv was cloned, overexpressed and purified. Crystals were obtained by the hanging-drop vapor-diffusion method and contained a 23 kDa GroEL1 fragment. A complete native data set was collected from a single frozen crystal that belonged to the orthorhombic space group P2(1)2(1)2, with unit-cell parameters a = 75.47, b = 78.67, c = 34.89 A, alpha = beta = gamma = 90 degrees , and diffracted to 2.2 A resolution on a home X-ray source. more...