Mycobacterium tuberculosispantothenate kinase: possible changes in location of ligands during enzyme action

Abstract

The crystal structures of complexes of Mycobacterium tuberculosis pantothenate kinase with the following ligands have been determined: (i) citrate; (ii) the nonhydrolysable ATP analogue AMPPCP and pantothenate (the initiation complex); (iii) ADP and phosphopantothenate resulting from phosphorylation of pantothenate by ATP in the crystal (the end complex); (iv) ATP and ADP, each with half occupancy, resulting from a quick soak of crystals in ATP (the intermediate complex); (v) CoA; (vi) ADP prepared by soaking and cocrystallization, which turned out to have identical structures, and (vii) ADP and pantothenate. Solution studies on CoA binding and catalytic activity have also been carried out. Unlike in the case of the homologous Escherichia coli enzyme, AMPPCP and ADP occupy different, though overlapping, locations in the respective complexes; the same is true of pantothenate in the initiation complex and phosphopantothenate in the end complex. The binding site of MtPanK is substantially preformed, while that of EcPanK exhibits considerable plasticity. The difference in the behaviour of the E. coli and M. tuberculosis enzymes could be explained in terms of changes in local structure resulting from substitutions. It is unusual for two homologous enzymes to exhibit such striking differences in action. Therefore, the results have to be treated with caution. However, the changes in the locations of ligands exhibited by M. tuberculosis pantothenate kinase are remarkable and novel.

Protein targets for structure-based anti-Mycobacterium tuberculosis drug discovery

“…Therefore, the four enzymes in pantothenate biosynthesis (Pan B-E) and lumazine synthase (LS) that catalyzes vitamin B5 and B2 synthesis, are characterized as attractive targets for anti-M. tuberculosis drug discovery (Cole et al, 2001;Sambandamurthy et al, 2002;Visca et al, 2002). Currently, several crystal structures are available and present the possibility for highly selective and sensitive anti-M. tuberculosis drug design (Cole et al, 2001;Chaudhuri et al, 2003;Eisenberg, 2003, 2006;Chetnani et al, 2009). …”

M. tuberculosis Pantothenate Kinase: Dual Substrate Specificity and Unusual Changes in Ligand Locations

“…7 However, unlike in the case of EcPanK, the structure observed in the MtPanK-CoA complex is retained in the complexes involving ADP and AMPPCP, another non-hydrolyzable analogue of ATP as well. 11 Furthermore, the locations of the nucleotide and the pantothenate are different in the ternary binitiationQ complex with AMPPCP and pantothenate and the bendQ complex involving ADP and phosphopantothenate. Thus, the enzyme action involves substantial movements of ligands in MtPanK unlike in EcPanK.…”

“…4,5 The structures of the enzyme in the CoA complex and in the complexes involving AMPPNP and ADP also show significant differences. 4,5 We have been exploring the structures and interactions of PanK from Mycobacterium tuberculosis (MtPanK) as part of a program in this laboratory [6][7][8][9][10][11][12] and an international effort involving structural studies on mycobacterial proteins. [13][14][15][16][17] The CoA complex of MtPanK has a structure very similar to that of the corresponding EcPanK complex.…”

Structural and Biochemical Characterization of Compounds Inhibiting Mycobacterium tuberculosis Pantothenate Kinase

“…4a). Residues Tyr-235 and Asn-277 are involved in pantothenate and phosphopantothenate binding (14), respectively, but here, the hydroxyl group of Tyr-235 forms a hydrogen bond to one nitrogen in the triazole ring, whereas the amide group of the side chain of Asn-277 hydrogen bonds to another (Fig. 4a) ring that maintains the protein-inhibitor hydrogen bonds but results in relatively large local changes in the enzyme.…”

“…It functions as a homodimer. MtPanK has been the target for extensive structural studies and has been crystallized in complex with its substrates, products, adenosine or guanosine cofactors, and feedback inhibitor CoA (13)(14)(15)(16). The MtPanK structures in this study represent the first complexes with engineered inhibitory compounds and provide a starting point for development of novel antibiotics targeting pathogens that are dependent on type I PanK.…”