Targeting IspD for Anti-infective and Herbicide Development: Exploring Its Role, Mechanism, and Structural Insights

Abstract

Antimicrobial resistance (AMR) and herbicide resistance pose threats to society, necessitating novel anti-infectives and herbicides exploiting untapped modes of action like inhibition of IspD, the third enzyme in the MEP pathway. The MEP pathway is essential for a wide variety of human pathogens, including Pseudomonas aeruginosa, Mycobacterium tuberculosis, and Plasmodium falciparum, as well as plants. Within the current perspective, we focused our attention on the third enzyme in this pathway, IspD, offering a comprehensive summary of the reported modes of inhibition and common trends, with the goal to inspire future research dedicated to this underexplored target. In addition, we included an overview of the history, catalytic mechanism, and structure of the enzyme to facilitate access to this attractive target.

Scheme 1. Overview of the MEP Pathway

Figure 1

IspD research from initial discovery to different homologues, mechanism, and inhibitors. methyl-D-erythritol phosphate (MEP)^,,

Scheme 2. Representation of the Proposed Reaction Mechanisms Catalyzed by IspD, Leading to the Formation of 4-Diphosphocytidyl-2C-methyl-d-erythritol (CDP-ME)

Figure 2

Crystal structure of Escherichia coli IspD with CTP in the active site (PDB accession code: 1I52). Visual representation of the two subunits. Green: larger subunit (residues 1–136 and 160–236); Blue: smaller subunit (residues 137–159).

Figure 3

Co-crystal structure of cytidine and CDP-ME in the active site of Escherichia coli IspD (PDB accession code: 1INI and 1I52); highly conserved residues (*) present in most IspD homologues; strictly conserved (^†) present in all IspD homologues. Top: overview of the interactions of the cytidine present in both crystal structures; middle: summary of the interactions of the triphosphate tail of CTP; bottom: interactions of the phosphate and MEP part of CDP-ME. Highly conserved residues (*): Pro-13, Ala-14, Ala-15, Gly-16, Arg-19, Ser-88, Ala-107, Ala-163. Strictly conserved residues (^†): Gly-18, Arg-20, Lys-27, Gly-82, Asp-106, Arg-109, Lys-213. Not conserved residues: Phe-17, Asp-83.

Figure 4

Overview of 2-phenyl benzo[d]isothiazol-3(2H)-one (BITZ) compounds and their IC₅₀ values against Plasmodium falciparum (Pf) and P. vivax (Pv) IspD, and growth inhibition against the Pf strain 3D7.

Scheme 3. Presumed Covalent Mechanism of 2-phenyl benzo[d]isothiazol-3(2H)-one (BITZ)

Figure 5

Overview of the enantiomers of MMV-008138 and their IC₅₀’s against Plasmodium falciparum (Pf) IspD; ^aref (36); ^bref (37).

Figure 6

Overview of the structural features key for the activity of the MMV-008138 chemotype.

Figure 7

New chemical classes derived from the MMV-008138 chemotype demonstrating promising growth inhibition of Plasmodium falciparum (Pf) strain Dd2; 8 ref (66); 9 ref (67).

Figure 8

MEPN₃ analogue of 2-C-methyl-d-erythritol 4-phosphate (MEP).

Figure 9

Crystal structure of Arabidopsis thaliana IspD with 11 in the allosteric pocket displaying the key interactions, which 11 is engaging with the allosteric pocket (PDB accession code: 2YC3). The allosteric pocket consists of the following residues: Leu-45, Arg-157, Val-161, Ala-202, Val-204, Gln-238, Val-239, Ile-240, Phe-249, Asp-262, Val-263 Ser-264, Ile-265, Val-266, and Val-273.

Figure 10

Initial hit (11) of the azolopyridine class with the points of interaction in the allosteric pocket highlighted.

Figure 11

Derivatives of the azolopyridine class designed to displace a water molecule inside the allosteric pocket.

Figure 12

Summary of the interactions between the pseudilines, Arabidopsis thaliana IspD, and Cd₂⁺.

Figure 13

Chemical structure of compounds 16-18, their activity against AtIspD and summary of the interactions with Arabidopsis thaliana IspD.

Figure 14

Top: initial pyrrolopyrazine, Arabidopsis thaliana (At); bottom: pyrrolopyrazine derivatives, Plasmodium falciparum (Pf).

Figure 15

Top: initial urea hit; bottom: promising urea compound. Activities against both Plasmodium falciparum (Pf) IspD and PfNF54 strain are displayed.

Figure 16

Structures of the initial hit and optimized compound of the biphenyl carboxylic acid, Plasmodium falciparum (Pf).

Figure 17

Structures of the initial hits of the aminobenzothiazole inhibitors, Arabidopsis thaliana (At).

Figure 18

2-C-Methyl-d-erythritol 4-phosphate (MEP) and desmethyl MEP.

Figure 19

Overview of the interactions of 4-diphosphocytidyl-2-C-methyl-d-erythritol (CDP-ME).

Figure 20

Overview fluoroalkyl phosphonyl analogues of 2-C-methyl-d-erythritol 4-phosphate (MEP).^,