Slow-Binding Inhibition of Mycobacterium tuberculosis Shikimate Kinase by Manzamine Alkaloids

Tuberculosis represents a significant public health crisis. There is an urgent need for novel molecular scaffolds against this pathogen. We screened a small library of marine-derived compounds against shikimate kinase from Mycobacterium tuberculosis ( Mt SK), a promising target for antitubercular drug development. Six manzamines previously shown to be active against M. tuberculosis were characterized as Mt SK inhibitors: manzamine A ( 1 ), 8-hydroxymanzamine A ( 2 ), manzamine E ( 3 ), manzamine F ( 4 ), 6-deoxymanzamine X ( 5 ), and 6-cyclohexamidomanzamine A ( 6 ). All six showed mixed noncompetitive inhibition of Mt SK. The lowest K I values were obtained for 6 across all Mt SK–substrate complexes. Time-dependent analyses revealed two-step, slow-binding inhibition. The behavior of 1 was typical; initial formation of an enzyme–inhibitor complex (EI) obeyed an apparent K I of ∼30 μM with forward ( k 5 ) and reverse ( k 6 ) rate constants for isomerization to an EI* complex of 0.18 and 0.08 min –1, respectively. In contrast, 6 showed a lower K I for the initial encounter complex (∼1.5 μM), substantially faster isomerization to EI* ( k 5 = 0.91 min –1 ), and slower back conversion of EI* to EI ( k 6 = 0.04 min –1 ). Thus, the overall inhibition constants, K I *, for 1 and 6 were 10 and 0.06 μM, respectively. These findings were consistent with docking predictions of a favorable binding mode and a second, less tightly bound pose for 6 at Mt SK. Our results suggest that manzamines, in particular 6, constitute a new scaffold from which drug candidates with novel mechanisms of action could be designed for the treatment of tuberculosis by targeting Mt SK.