The structure of glucose‐1‐phosphate thymidylyltransferase from Mycobacterium tuberculosis reveals the location of an essential magnesium ion in the RmlA‐type enzymes

Abstract

Tuberculosis, caused by the bacterium Mycobacterium tuberculosis, continues to be a major threat to populations worldwide. Whereas the disease is treatable, the drug regimen is arduous at best with the use of four antimicrobials over a six-month period. There is clearly a pressing need for the development of new therapeutics. One potential target for structure-based drug design is the enzyme RmlA, a glucose-1-phosphate thymidylyltransferase. This enzyme catalyzes the first step in the biosynthesis of l-rhamnose, which is a deoxysugar critical for the integrity of the bacterium's cell wall. Here, we report the X-ray structures of M. tuberculosis RmlA in complex with either dTTP or dTDP-glucose to 1.6 Å and 1.85 Å resolution, respectively. In the RmlA/dTTP complex, two magnesium ions were observed binding to the nucleotide, both ligated in octahedral coordination spheres. In the RmlA/dTDP-glucose complex, only a single magnesium ion was observed. Importantly, for RmlA-type enzymes with known three-dimensional structures, not one model shows the position of the magnesium ion bound to the nucleotide-linked sugar. As such, this investigation represents the first direct observation of the manner in which a magnesium ion is coordinated to the RmlA product and thus has important ramifications for structure-based drug design. In the past, molecular modeling procedures have been employed to derive a three-dimensional model of the M. tuberculosis RmlA for drug design. The X-ray structures presented herein provide a superior molecular scaffold for such endeavors in the treatment of one of the world's deadliest diseases.

Pangenome Analysis of Mycobacterium tuberculosis Reveals Core-Drug Targets and Screening of Promising Lead Compounds for Drug Discovery

“…The present study also prioritized drug target glucose-1-phosphate thymidylyltransferase. The protein is involved in three metabolic pathways namely, polyketide sugar unit biosynthesis, streptomycin biosynthesis, and the nucleotide sugars metabolism pathway [ 36 , 37 , 38 ]. All these pathways are crucial for survival of M. tuberculosis and elaborate on the relevance of glucose-1-phosphate thymidylyltransferase as a drug target.…”

“…These ligands are 2′deoxy-thymidine-5′-diphospho-alpha- d -glucose, uridine diphosphate glucose, 2′-deoxy-thymidine-beta- l -rhamnose, thymidine-5′-triphosphate, and citicoline, which have good binding affinity with metabolic pathway proteins especially glucose-1-phosphate thymidylyltransferase. The protein is involved in three metabolic pathways namely, polyketide sugar unit biosynthesis, streptomycin biosynthesis, and nucleotide sugars metabolism pathway [ 36 , 37 , 38 ]. All these pathways are crucial for survival of M. tuberculosis and shortlisted repurposed drugs have higher affinity with the enzyme, i.e., glucose-1-phosphate thymidylyltransferase.…”

The Mycobacterium tuberculosis complex has a pathway for the biosynthesis of 4‐formamido‐4,6‐dideoxy‐d‐glucose

“…There is an enzyme in M. tuberculosis , encoded by the rv0334 gene that catalyzes such a reaction. It is specifically involved in the biosynthesis of l ‐rhamnose, and its three‐dimensional structure has now been determined to high resolution . In the second step of the pathway, dTDP‐ d ‐glucose is dehydrated by the action of a 4,6‐dehydratase to yield dTDP‐4‐keto‐6‐deoxy‐ d ‐glucose.…”

“…It is specifically involved in the biosynthesis of L-rhamnose, 16 and its three-dimensional structure has now been determined to high resolution. 17 In the second step of the pathway, dTDP-D-glucose is dehydrated by the action of a 4,6-dehydratase to yield dTDP-4-keto-6deoxy-D-glucose. Again, this reaction is also required for the production of L-rhamnose in M. tuberculosis, and not surprisingly the gene encoding for an enzyme with such activity has been identified, namely rv3464.…”

Data_Sheet_1.PDF

“…Over the past few decades, many Rml enzymes from different prokaryotes have been reported (Graninger et al, 2002;Han et al, 2007;Steiner et al, 2008;van der Beek et al, 2019). Most of those works focused on the biological roles of these enzymes and their potential as drug targets for the treatment of pathogenic bacteria, such as Mycobacteria tuberculosis H37Rv (Brown et al, 2017), Streptococcus pyogenes 5,448 (van der Beek et al, 2019), Bacillus anthracis str. Ames (Gokey et al, 2018), etc.…”