Discovery, Synthesis and Evaluation of a Ketol‐Acid Reductoisomerase Inhibitor

Bayaraa, Tenuun; Kurz, Julia L.; Patel, Khushboo; Hussein, Waleed M.; Bilyj, Jessica K.; West, Nicholas P.; Schenk, Gerhard; McGeary, Ross P.; Guddat, L.W.

doi:10.1002/chem.202000899

Cited by 19 publications

(23 citation statements)

References 37 publications

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“…Sa KARI was chosen here because, in contrast to M tKARI, it readily crystallized with a variety of inhibitors [7b,c] . Importantly, the active sites of the two enzymes are virtually identical [12, 15] .…”

Section: Resultsmentioning

confidence: 99%

“…Sa KARI was expressed and purified as described previously [7c] . Crystals were formed by using the hanging drop method (3 μL drops) in VDX crystallization plates.…”

Section: Methodsmentioning

confidence: 99%

“…Despite multiple attempts at crystallization, no other structures for Mt KARI in complex with inhibitors; substrate analogues; or its cofactor, NADPH, have been determined. However, Staphylococcus aureus KARI ( Sa KARI), the overall sequence of which, especially in the active site, is highly homologous to that of Mt KARI, readily crystallizes in the presence of NADPH and several transition‐state analogues [7b,c] …”

Section: Introductionmentioning

confidence: 99%

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Discovery of a Pyrimidinedione Derivative with Potent Inhibitory Activity against Mycobacterium tuberculosis Ketol–Acid Reductoisomerase

Lin

Kurz

Patel

et al. 2021

Chemistry A European J

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New drugs aimed at novel targets are urgently needed to combat the increasing rate of drug‐resistant tuberculosis (TB). Herein, the National Cancer Institute Developmental Therapeutic Program (NCI‐DTP) chemical library was screened against a promising new target, ketol–acid reductoisomerase (KARI), the second enzyme in the branched‐chain amino acid (BCAA) biosynthesis pathway. From this library, 6‐hydroxy‐2‐methylthiazolo[4,5‐d]pyrimidine‐5,7(4H,6H)‐dione (NSC116565) was identified as a potent time‐dependent inhibitor of Mycobacterium tuberculosis (Mt) KARI with a Ki of 95.4 nm. Isothermal titration calorimetry studies showed that this inhibitor bound to MtKARI in the presence and absence of the cofactor, nicotinamide adenine dinucleotide phosphate (NADPH), which was confirmed by crystal structures of the compound in complex with closely related Staphylococcus aureus KARI. It is also shown that NSC116565 inhibits the growth of H37Ra and H37Rv strains of Mt with MIC50 values of 2.93 and 6.06 μm, respectively. These results further validate KARI as a TB drug target and show that NSC116565 is a promising lead for anti‐TB drug development.

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Section: Resultsmentioning

confidence: 99%

“…Sa KARI was expressed and purified as described previously [7c] . Crystals were formed by using the hanging drop method (3 μL drops) in VDX crystallization plates.…”

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Discovery of a Pyrimidinedione Derivative with Potent Inhibitory Activity against Mycobacterium tuberculosis Ketol–Acid Reductoisomerase

Lin

Kurz

Patel

et al. 2021

Chemistry A European J

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“… 11 KARI has also attracted considerable attention as an anti-TB drug target. 12 KARIs are bifunctional enzymes that require Mg 2+ as cofactor in their first half isomerization reaction and NADH or NADPH as reducing agents in the second half reduction reaction. 13 a Previous studies demonstrated that HMKB 3 is the transit intermediate catalysed by the KARI enzyme (EcoKARI) from E. coli ( Fig.…”

Section: Introductionmentioning

confidence: 99%

Characterization of a class II ketol-acid reductoisomerase from Mycobacterium tuberculosis

et al. 2022

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“…The enzyme is therefore necessary for amino acid prototrophy. Because of its importance in bacteria, plants and fungi, but its absence in metazoans, the enzyme receives increasing attention for the design of antibiotics, herbicides and fungicides [2][3][4]. On the other hand, branched-chain amino acids are valuable compounds for the food, cosmetic and pharmaceutical industries, with a constantly increasing global market (e.g., L-valine: average annual increase rate >5% [5,6]).…”

Section: Introductionmentioning

confidence: 99%

Structural Rearrangements of a Dodecameric Ketol-Acid Reductoisomerase Isolated from a Marine Thermophilic Methanogen

et al. 2021

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Ketol-acid reductoisomerase (KARI) orchestrates the biosynthesis of branched-chain amino acids, an elementary reaction in prototrophic organisms as well as a valuable process in biotechnology. Bacterial KARIs belonging to class I organise as dimers or dodecamers and were intensively studied to understand their remarkable specificity towards NADH or NADPH, but also to develop antibiotics. Here, we present the first structural study on a KARI natively isolated from a methanogenic archaea. The dodecameric structure of 0.44-MDa was obtained in two different conformations, an open and close state refined to a resolution of 2.2-Å and 2.1-Å, respectively. These structures illustrate the conformational movement required for substrate and coenzyme binding. While the close state presents the complete NADP bound in front of a partially occupied Mg2+-site, the Mg2+-free open state contains a tartrate at the nicotinamide location and a bound NADP with the adenine-nicotinamide protruding out of the active site. Structural comparisons show a very high conservation of the active site environment and detailed analyses point towards few specific residues required for the dodecamerisation. These residues are not conserved in other dodecameric KARIs that stabilise their trimeric interface differently, suggesting that dodecamerisation, the cellular role of which is still unknown, might have occurred several times in the evolution of KARIs.

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Discovery, Synthesis and Evaluation of a Ketol‐Acid Reductoisomerase Inhibitor

Abstract: This is the author manuscript accepted for publication and has undergone full peer review but has not been through the copyediting, typesetting, pagination and proofreading process, which may lead to differences between this version and the Version of Record.

Cited by 19 publications

References 37 publications

Discovery of a Pyrimidinedione Derivative with Potent Inhibitory Activity against Mycobacterium tuberculosis Ketol–Acid Reductoisomerase

Discovery of a Pyrimidinedione Derivative with Potent Inhibitory Activity against Mycobacterium tuberculosis Ketol–Acid Reductoisomerase

Characterization of a class II ketol-acid reductoisomerase from Mycobacterium tuberculosis

Structural Rearrangements of a Dodecameric Ketol-Acid Reductoisomerase Isolated from a Marine Thermophilic Methanogen

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