A comparative biochemical and structural analysis of the intracellular chorismate mutase (Rv0948c) from Mycobacterium tuberculosis H₃₇R_v and the secreted chorismate mutase (y2828) from Yersinia pestis

Abstract

The Rv0948c gene from Mycobacterium tuberculosis H37Rv encodes a 90 amino acid protein as the natural gene product with chorismate mutase (CM) activity. The protein, 90‐MtCM, exhibits Michaelis–Menten kinetics with a kcat of 5.5 ± 0.2 s−1 and a Km of 1500 ± 100 μm at 37 °C and pH 7.5. The 2.0 Å X‐ray structure shows that 90‐MtCM is an all α‐helical homodimer (Protein Data Bank ID: 2QBV) with the topology of Escherichia coli CM (EcCM), and that both protomers contribute to each catalytic site. Superimposition onto the structure of EcCM and the sequence alignment shows that the C‐terminus helix 3 is shortened. The absence of two residues in the active site of 90‐MtCM corresponding to Ser84 and Gln88 of EcCM appears to be one reason for the low kcat. Hence, 90‐MtCM belongs to a subfamily of α‐helical AroQ CMs termed AroQδ. The CM gene (y2828) from Yersinia pestis encodes a 186 amino acid protein with an N‐terminal signal peptide that directs the protein to the periplasm. The mature protein, *YpCM, exhibits Michaelis–Menten kinetics with a kcat of 70 ± 5 s−1 and Km of 500 ± 50 μm at 37 °C and pH 7.5. The 2.1 Å X‐ray structure shows that *YpCM is an all α‐helical protein, and functions as a homodimer, and that each protomer has an independent catalytic unit (Protein Data Bank ID: 2GBB). *YpCM belongs to the AroQγ class of CMs, and is similar to the secreted CM (Rv1885c, *MtCM) from M. tuberculosis.

Complex Formation between Two Biosynthetic Enzymes Modifies the Allosteric Regulatory Properties of Both

“…Because no significant conformational change of the protein is observed on inhibitor binding, except in the near vicinity of the Phe binding site, inhibition of MtuDAH7PS is consistent with a change in protein dynamics, causing altered substrate binding (12). The genome of M. tuberculosis encodes two CM proteins (13,14). One is highly active and secreted into the periplasmic compartment, and its biological role is yet to be determined (14).…”

Structure of Chorismate Mutase-like Domain of DAHPS from Bacillus subtilis Complexed with Novel Inhibitor Reveals Conformational Plasticity of Active Site

“…1D), and similarly in chain C, and chain D. The citrate molecule also makes additional interactions with the residues Arg27, Arg50 and Phe79 via water bridge formation. It has been previously reported that the citrate inhibits the 90- Mt CM protein 34 .…”

“…Residues Phe57′, Ala59′ and Leu115 are also part of the active site 34 . The Bs CM_2 active site consists of similar residues.…”

FREQUENCY OF MUTATION IN ISONIAZID RESISTANT ISOLATES OF Mycobacterium tuberculosis COMPLEX FROM WESTERN MAHARASHTRA INDIA

“…Samples were processed for ZN staining before and after Decontamination of the sample with NALC-NaOH method. [6] Samples were processed for two molecular methods Line Probe Assay (LiPA), [7] (Hain Life sciences GmbH, Nerhren, Germany) and Sanger Sequencing (3130 genetic analyzer, Applied Biosystems) [8]. Study was done on monodrug and multidrug resistant strains of MTB complex.…”