6-Arylpyrido[2,3-d]pyrimidines as Novel ATP-Competitive Inhibitors of Bacterial D-Alanine:D-Alanine Ligase (original) (raw)
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ATP competitive inhibitors of d-alanine–d-alanine ligase based on protein kinase inhibitor scaffolds
Bioorganic & Medicinal Chemistry, 2009
D D-Alanine-D D-alanine ligase (DDl) is an essential enzyme in bacterial cell wall biosynthesis and an important target for developing new antibiotics. Here, we describe a new approach to identify new inhibitor scaffolds for DDl based on similarity in the ATP binding region of different kinases and DDl. After an initial screening of several protein kinase inhibitors, we found that the Brutons's tyrosine kinase inhibitor LFM-A13, an analog of the Leflunomide metabolite A771726, inhibits DDl with a K i of 185 lM. A series of malononitrilamide and salicylamide derivatives of LFM-A13 has been synthesized to confirm the validity of this scaffold as an inhibitor of DDl.
Medicinal Chemistry Research, 2014
D-Alanine:D-alanine ligase (Ddl) catalyzes the ATP-driven ligation of two D-alanine (D-Ala) molecules resulting in the formation of D-alanyl:D-alanine dipeptide. Inhibition of Ddl prevents bacterial growth, which makes this enzyme an attractive and viable target for searching effective antimicrobial drugs. Current study aimed at the discovery of potent inhibitors against Strepotococcus sanguinis SK36 Ddl (SsDdl). Using comparative molecular modeling approach, models for SsDdl were constructed by MODELLER and other web servers. The model with best stereo-chemical profile was further studied for structure function relationship by performing docking with seventy nine inhibitors. The results demonstrated the model generated via MODELLER was the best among all predicted SsDdl structures. Docking analysis revealed that compounds 73 and 61, obtained from pyridopyrimidine scaffolds yielded highest GOLD scores and exhibited consistently better binding interactions. The final compounds exhibit reasonable SsDdl inhibitory activity and can be further employed to design derivatives with customized activities.
Discovery of New Inhibitors of d -Alanine: d -Alanine Ligase by Structure-Based Virtual Screening †
Journal of Medicinal Chemistry, 2008
The terminal dipeptide, D-Ala-D-Ala, of the peptidoglycan precursor UDPMurNAc-pentapetide is a crucial building block involved in peptidoglycan cross-linking. It is synthesized in the bacterial cytoplasm by the enzyme D-alanine:D-alanine ligase (Ddl). Structure-based virtual screening of the NCI diversity set of almost 2000 compounds was performed with a DdlB isoform from Escherichia coli using the computational tool AutoDock 4.0. The 130 best-ranked compounds from this screen were tested in an in vitro assay for their inhibition of E. coli DdlB. Three compounds were identified that inhibit the enzyme with K i values in micromolar range. Two of these also have promising antibacterial activities against Gram-positive and Gramnegative bacteria.
Inhibitors of ATP Synthase as New Antibacterial Candidates
Antibiotics
ATP, the power of all cellular functions, is constantly used and produced by cells. The enzyme called ATP synthase is the energy factory in all cells, which produces ATP by adding inorganic phosphate (Pi) to ADP. It is found in the inner, thylakoid and plasma membranes of mitochondria, chloroplasts and bacteria, respectively. Bacterial ATP synthases have been the subject of multiple studies for decades, since they can be genetically manipulated. With the emergence of antibiotic resistance, many combinations of antibiotics with other compounds that enhance the effect of these antibiotics have been proposed as approaches to limit the spread of antibiotic-resistant bacteria. ATP synthase inhibitors, such as resveratrol, venturicidin A, bedaquiline, tomatidine, piceatannol, oligomycin A and N,N-dicyclohexylcarbodiimide were the starting point of these combinations. However, each of these inhibitors target ATP synthase differently, and their co-administration with antibiotics increases t...
Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics, 2004
UDP-N-acetylmuramyl-L-alanine ligase (MurC) is an essential bacterial enzyme involved in peptidoglycan biosynthesis and a target for the discovery of novel antibacterial agents. As a result of a high-throughput screen (HTS) against a chemical library for inhibitors of MurC, a series of benzofuran acyl-sulfonamides was identified as potential leads. One of these compounds, Compound A, inhibited Escherichia coli MurC with an IC 50 of 2.3 AM. Compound A exhibited time-dependent, partially reversible inhibition of E. coli MurC. Kinetic studies revealed a mode of inhibition consistent with the compound acting competitively with the MurC substrates ATP and UDP-N-acetyl-muramic acid (UNAM) with a K i of 4.5 AM against ATP and 6.3 AM against UNAM. Fluorescence binding experiments yielded a K d of 3.1 AM for the compound binding to MurC. Compound A also exhibited high-affinity binding to bovine serum albumin (BSA) as evidenced by a severe reduction in MurC inhibition upon addition of BSA. This finding is consistent with the high lipophilicity of the compound. Advancement of this compound series for further drug development will require reduction of albumin binding. D Abbreviations: AMP-PCP, h,g-methyleneadenosine 5V -triphosphate; BSA, bovine serum albumin; DLS, dynamic light scattering; DMSO, dimethyl sulfoxide; HTS, high-throughput screen; IC 50 , concentration of inhibitor which provides 50% inhibition; log D, octanol to water partition coefficient at pH 7.4 in phosphate buffer; log P, octanol to water partition
Ellipticines and 9-acridinylamines as inhibitors of d-alanine:d-alanine ligase
Bioorganic & Medicinal Chemistry, 2011
a b s t r a c t D-Alanine:D-alanine ligase (Ddl), an intracellular bacterial enzyme essential for cell wall biosynthesis, is an attractive target for development of novel antimicrobial drugs. This study focused on an extensive evaluation of two families of Ddl inhibitors encountered in our previous research. New members of both families were obtained through similarity search and synthesis. Ellipticines and 9-acridinylamines were both found to possess inhibitory activity against Ddl from Escherichia coli and antimicrobial activity against E. coli and Staphylococcus aureus. Ellipticines with a quaternary methylpyridinium moiety were the most potent among all studied compounds, with MIC values as low as 2 mg/L in strains with intact efflux mechanisms. Antimicrobial activity of the studied compounds was connected to membrane damage, making their development as antibacterial drug candidates unlikely unless analogues devoid of this nonspecific effect can be discovered.
Antimicrobial Agents and Chemotherapy, 2011
D-alanine:D-alanine ligase (EC 6.3.2.4; Ddl) catalyzes the ATP-driven ligation of two D-alanine (D-Ala) molecules to form the D-alanyl:D-alanine dipeptide. This molecule is a key building block in peptidoglycan biosynthesis, making Ddl an attractive target for drug development. D-Cycloserine (DCS), an analog of D-Ala and a prototype Ddl inhibitor, has shown promise for the treatment of tuberculosis. Here, we report the crystal structure of Mycobacterium tuberculosis Ddl at a resolution of 2.1 Å. This structure indicates that Ddl is a dimer and consists of three discrete domains; the ligand binding cavity is at the intersection of all three domains and conjoined by several loop regions. The M. tuberculosis apo Ddl structure shows a novel conformation that has not yet been observed in Ddl enzymes from other species. The nucleotide and D-alanine binding pockets are flexible, requiring significant structural rearrangement of the bordering regions for entry and binding of both ATP and D-Ala molecules. Solution affinity and kinetic studies showed that DCS interacts with Ddl in a manner similar to that observed for D-Ala. Each ligand binds to two binding sites that have significant differences in affinity, with the first binding site exhibiting high affinity. DCS inhibits the enzyme, with a 50% inhibitory concentration (IC(50)) of 0.37 mM under standard assay conditions, implicating a preferential and weak inhibition at the second, lower-affinity binding site. Moreover, DCS binding is tighter at higher ATP concentrations. The crystal structure illustrates potential drugable sites that may result in the development of more-effective Ddl inhibitors.
Biochemistry, 1988
In Salmonella typhimurium, D-a1anine:D-alanine ligase (ADP) (EC 6.3.2.4) is the second enzyme in the three enzyme palanine branch pathway of peptidoglycan biosynthesis. The interaction of this enzyme with a possible transition-state analogue, the (aminoalky1)phosphinate D-3-[ (l-aminoethyl)phosphinyl]-2heptylpropionic acid [Parsons et al. (1987) Abstracts of Papers, 193rd National Meeting of the American Chemical Society, Denver, CO, MEDI 63, American Chemical Society, Washington, DC] , has been studied. This compound is a potent active site directed inhibitor and is competitive with D-alanine (Ki = 1.2 pM); it exhibits time-dependent inhibition in the presence of ATP. Kinetic analysis revealed a rapid onset of steady-state inhibition (k,, = 1.35 X lo4 M-' s-' ) followed by slow dissociation of inhibitory complex(es) with a half-life of 8.2 h. The inhibitory complex was shown to consist of E-.I--ATP in equilibrium with E--I, Pi, and ADP. Similar time-dependent inhibition was also observed with D-( 1-aminoethy1)phosphonic acid (D-Ala-P) (Ki = 0.5 mM; k,, = 27 M-' s-l; t l I 2 for regain = 1.73 min) but not with D-(1-amino-ethy1)phosphinic acid, which behaved as a simple competitive inhibitor (Ki = 0.4 mM). The mechanism of inhibition is discussed in the light of the precedents of glutamine synthase inhibition by methionine sulfoximine and phosphinothricin.
Inhibition of D-Ala:D-Ala ligase through a phosphorylated form of the antibiotic D-cycloserine
Nature communications, 2017
D-cycloserine is an antibiotic which targets sequential bacterial cell wall peptidoglycan biosynthesis enzymes: alanine racemase and D-alanine:D-alanine ligase. By a combination of structural, chemical and mechanistic studies here we show that the inhibition of D-alanine:D-alanine ligase by the antibiotic D-cycloserine proceeds via a distinct phosphorylated form of the drug. This mechanistic insight reveals a bimodal mechanism of action for a single antibiotic on different enzyme targets and has significance for the design of future inhibitor molecules based on this chemical structure.