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Logo of jbcThe Journal of Biological Chemistry
 
J Biol Chem. 2016 July 1; 291(27): 14386–14387.
PMCID: PMC4933192

Structural and biochemical insight into the mechanism of Rv2837c from Mycobacterium tuberculosis as a c-di-NMP phosphodiesterase.

VOLUME 291 (2016) PAGES 3668–3681

Based on the reprocessed diffraction data for Rv2837c crystal, a better model and electron density was obtained at a little higher resolution of 2.3 Å (Table 1). The new electron density shows that the two protein molecules in the asymmetric unit are in different states. In molecule A, the FoFc map clearly indicates the existence of a catalytic water molecule between the two Mn2+ ions and pApA-like molecule. In molecule B, water molecule is not observed, and the FoFc map suggests two AMPs (Fig. 10). As a precaution, both pApA and AMP were fitted in the active site of each protein molecule and refined. The result clearly shows that two AMPs should be predominant in molecule B and that a pApA fits into the electron density better than two AMPs in molecule A. It also indicates that AMPs are not compatible with the catalytic water molecule. Consequently, we hypothesize that even though molecule A contains a mixture of pApA and AMP, pApA still outweighs AMP (Fig. 11). Thus, in the final model, a pApA is placed in the active site of molecule A, and two AMPs are placed in the active site of molecule B. That means our structure not only catches a snapshot in the middle of catalysis, but it also shows the last moment of the catalysis before the product leaves the active site. It is also confirmed that the new structure is still supportive to the mechanism previously proposed in the paper.

TABLE 1
X-ray diffraction data collection and refinement statistics
FIGURE 10.
The pApA and AMP were placed in the active sites of Rv2837c dimer according to the FoFc map that is contoured at the 3σ and 2σ level, respectively. A, stereoview of the pApA at the active site of molecule A. B, stereoview of ...
FIGURE 11.
Electron density for pApA and AMP of 2FoFc map contoured at the 1.0σ level. A, stereoview of pApA at the active site of molecule A. B, stereoview of two AMPs at the active site of molecule A. C, stereoview of two AMPs at the active ...

The atomic coordinates and structure factors (code 5JJU; Rv2837c-pApA/AMP) have been deposited in the Protein Data Bank (http://wwpdb.org/).


Articles from The Journal of Biological Chemistry are provided here courtesy of American Society for Biochemistry and Molecular Biology