Increase In Inhibitory Activity Research Articles

Inhibition of monoamine oxidases and neuroprotective effects: chalcones vs. chromones.

Eighteen compounds derived from two sub-series, (HC1-HC9) and (HF1-HF9), were synthesized and evaluated for their inhibitory activities against monoamine oxidase (MAO). HC (chalcone) series showed higher inhibitory activity against MAO-B than against MAO-A, whereas the HF (chromone) series showed reversed inhibitory activity. Compound HC4 most potently inhibited MAO-B with an IC50 value of 0.040μM, followed by HC3 (IC50 = 0.049μM), while compound HF4 most potently inhibited MAO-A (IC50 = 0.046μM), followed by HF2 (IC50 = 0.075μM). The selectivity index (SI) values of HC4 and HF4 were 50.40 and 0.59, respectively. Structurally, HC4 (4-OC2H5 in B-ring) showed higher MAO-B inhibition than other derivatives, suggesting that the -OC2H5 substitution of the 4-position in the B-ring contributes to the increase of MAO-B inhibition, especially -OC2H5 (HC4) > -OCH3 (HC3) > -F (HC7) > -CH3 (HC2) > -Br (HC8) > -H (HC1) in order. In MAO-A inhibition, the substituent 4-OC2H5 in the B-ring of HF4 contributed to an increase in inhibitory activity, followed by -CH3 (HF2), -F (HF7), -Br (HF8), -OCH3 (HF3), and-H (HF1). In the enzyme kinetics and reversibility study, the Ki value of HC4 for MAO-B was 0.035 ± 0.005μM, and that of HF4 for MAO-A was 0.035 ± 0.005μM, and both were reversible competitive inhibitors. We confirmed that HC4 and HF4 significantly ameliorated rotenone-induced neurotoxicity, as evidenced by the reactive oxygen species and superoxide dismutase assays. This study also supports the significant effect of HC4 and HF4 on mitochondrial membrane potential in rotenone-induced toxicity. A lead molecule was used for molecular docking and dynamic simulation studies. These results show that HC4 is a potent selective MAO-B inhibitor and HF4 is a potent MAO-A inhibitor, suggesting that both compounds can be used as treatment agents for neurological disorders.

Arylsulfonamido-alkyl-sulfamates act as inhibitors of bovine carbonic anhydrase II

A small library of arylsulfonamido-alkyl sulfamates was prepared by a two-step synthesis from readily available starting materials. The compounds were tested for their ability to inhibit bovine carbonic anhydrase II. Several of them were found as good competitive inhibitors holding Ki values as low as Ki = 0.9 μM (compound 47b). The activity was influenced by the substitution pattern of the arylsulfonamide moiety as well as the length of the spacer to the distal sulfamate group. Molecular docking studies were used to substantiate these findings. For the aryl-substituted analogues, the increase in inhibitory activity for compounds with a shorter spacer can be explained by stabilization via aromatic π-interactions. For the cyclopropyl or methylsulfonyl substituted analogues, their inhibitory activity can be attributed to their reduced steric hindrance. These results provide a basis for designing effective CA II inhibitors.

Synthesis and Activity Evaluation of Vinpocetine-Derived Indole Alkaloids.

This study focuses on the synthesis of novel vinpocetine derivatives (2-25) and their biological evaluation. The chemical structures of the synthesized compounds were fully characterized using techniques such as 1H NMR, 13C NMR, and HRMS. The inhibitory activity of the synthesized compounds on PDE1A was evaluated, and the results revealed that compounds 3, 4, 5, 12, 14, 21, and 25 exhibited superior inhibitory activity compared to vinpocetine. Compound 4, with a para-methylphenyl substitution, showed a 5-fold improvement in inhibitory activity with an IC50 value of 3.53 ± 0.25 μM. Additionally, compound 25, with 3-chlorothiazole substitution, displayed an 8-fold increase in inhibitory activity compared to vinpocetine (IC50 = 2.08 ± 0.16 μM). Molecular docking studies were conducted to understand the binding models of compounds 4 and 25 within the active site of PDE1A. The molecular docking study revealed additional binding interactions, such as π-π stacking and hydrogen bonding, contributing to the enhanced inhibitory activity and stability of the ligand-protein complexes. Overall, the synthesized vinpocetine derivatives demonstrated promising inhibitory activity on PDE1A, and the molecular docking studies provided insights into their binding modes, supporting further development of these compounds as potential candidates for drug research and development.

Excessive excitability of inhibitory cortical circuit and disturbance of ballistic targeting movement in degenerative cerebellar ataxia

This study aimed to investigate abnormalities in inhibitory cortical excitability and motor control during ballistic-targeting movements in individuals with degenerative cerebellar ataxia (DCA). Sixteen participants took part in the study (DCA group [n = 8] and healthy group [n = 8]). The resting motor-threshold and cortical silent period (cSP) were measured in the right-hand muscle using transcranial magnetic stimulation over the left primary motor cortex. Moreover, the performance of the ballistic-targeting task with right wrist movements was measured. The Scale for the Assessment and Rating of Ataxia was used to evaluate the severity of ataxia. The results indicated that the cSP was significantly longer in participants with DCA compared to that in healthy controls. However, there was no correlation between cSP and severity of ataxia. Furthermore, cSP was linked to the ballistic-targeting task performance in healthy participants but not in participants with DCA. These findings suggest that there is excessive activity in the gamma-aminobutyric acid-mediated cortical inhibitory circuit in individuals with DCA. However, this increase in inhibitory activity not only fails to contribute to the control of ballistic-targeting movement but also shows no correlation with the severity of ataxia. These imply that increased excitability in inhibitory cortical circuits in the DCA may not contribute the motor control as much as it does in healthy older adults under limitations associated with a small sample size. The study's results contribute to our understanding of motor control abnormalities in people with DCA and provide potential evidence for further research in this area.

Precision spinal gene delivery-induced functional switch in nociceptive neurons reverses neuropathic pain

Second-order spinal cord excitatory neurons play a key role in spinal processing and transmission of pain signals to the brain. Exogenously induced change in developmentally imprinted excitatory neurotransmitter phenotypes of these neurons to inhibitory has not yet been achieved. Here, we use a subpial dorsal horn-targeted delivery of AAV (adeno-associated virus) vector(s) encoding GABA (gamma-aminobutyric acid) synthesizing-releasing inhibitory machinery in mice with neuropathic pain. Treated animals showed a progressive and complete reversal of neuropathic pain (tactile and brush-evoked pain behavior) that persisted for a minimum of 2.5months post-treatment. The mechanism of this treatment effect results from the switch of excitatory to preferential inhibitory neurotransmitter phenotype in dorsal horn nociceptive neurons and a resulting increase in inhibitory activity in regional spinal circuitry after peripheral nociceptive stimulation. No detectable side effects (e.g., sedation, motor weakness, loss of normal sensation) were seen between 2 and 13months post-treatment in naive adult mice, pigs, and non-human primates. The use of this treatment approach may represent a potent and safe treatment modality in patients suffering from spinal cord or peripheral nerve injury-induced neuropathic pain.

Amorphous Inclusion Complexes: Molecular Interactions of Hesperidin and Hesperetin with HP-Β-CD and Their Biological Effects.

This study aimed at obtaining hesperidin (Hed) and hesperetin (Het) systems with HP-β-CD by means of the solvent evaporation method. The produced systems were identified using infrared spectroscopy (FT-IR), X-ray powder diffraction (XRPD), and differential scanning calorimetry (DSC). Moreover, in silico docking and molecular dynamics studies were performed to assess the most preferable site of interactions between tested compounds and HP-β-CD. The changes of physicochemical properties (solubility, dissolution rate, and permeability) were determined chromatographically. The impact of modification on biological activity was tested in an antioxidant study as well as with regards to inhibition of enzymes important in pathogenesis of neurodegenerative diseases. The results indicated improvement in solubility over 1000 and 2000 times for Hed and Het, respectively. Permeability studies revealed that Hed has difficulties in crossing biological membranes, in contrast with Het, which can be considered to be well absorbed. The improved physicochemical properties influenced the biological activity in a positive manner by the increase in inhibitory activity on the DPPH radical and cholinoesterases. To conclude the use of HP-β-CD as a carrier in the formation of an amorphous inclusion complex seems to be a promising approach to improve the biological activity and bioavailability of Hed and Het.

Azo-incorporating Increases Inhibitory Activity of Succinate Dehydrogenase

Diversity of pesticide discovery provided a solution to resistance. Here, we presented a strategy of azo-incorporating to promote the diverse developments of fungicide. A series of novel fungicides were synthesized by incorporating azobenzene derivatives into fluxapyroxad. Much better in vitro fungicidal activity increases for compound 9d were observed compared to the positive control, fluxapyroxad against Botrytis cinerea and Rhizoctonia solani. Compound 9d (IC50 = 0.03 μM) also had a great enzyme-inhibiting activity increase toward succinate dehydrogenase in comparison with fluxapyroxad (IC50 = 4.40 μM). A comparatively equivalent biological activity was observed between compounds 8a and 9d. SEM analysis helped us to observe clearly the morphology of the fungi before and after active ingredient delivery. Our results of molecular docking analysis, fluorescence quenching analysis, and enzymatic assays demonstrated that compound 8a and 9d act on SDH. An increase in inhibitory activity could be occurring after incorporation of azobenzene, which provided a new strategy for molecular design in pesticide discovery.

Synthesis of AgNPs and Ag@MoS2 nanocomposites by dracocephalum kotschyi aqueous extract and their antiacetylcholinesterase activities

ABSTRACT Green synthesis of metal nanoparticles and nanocomposites offers numerous advantages over conventional methods. The present study deals with the synthesis of Ag nanoparticles and Ag@MoS2 nanocomposites by Dracocephalum kotschyi extract as a reductant and a stabilising agent. The influence of extract volume, reaction temperature, contact time and pH on the synthesis of d-AgNPs was investigated. Further, the modified MoS2 using D. kotschyi extract (d-MoS2) could deposit Ag+ ions onto the MoS2 nanosheets surface, reduce the ions to AgNPs, and stabilise the reduced particles effectively. The synthesised nanostructures were characterised by XRD, SEM-EDS, TEM, Zeta potential and FT-IR analysis. Ag nanoparticles are spherical with diameter of 58–75 nm for d-AgNPs and 35–55 nm for Ag@d-MoS2, respectively. The d-AgNPs exhibited high activity in the inhibition of acetylcholinesterase. Also, loading of AgNPs on MoS2 nanosheets resulted in the increase of inhibitory activity against acetylcholinesterase compared to MoS2 nanosheets.

Design of Tripeptides as a Competitive Inhibitor for HMG-CoA Reductase

This study presents a simple approach in design of tripeptides as a competitive inhibitor for 3-hydroxy-3-methylglutaryl CoA reductase (HMGR). HMGR is a major rate limiting enzyme in cholesterol biosynthesis and therefore constitutes the target enzyme of an extensive research to discover new compounds focused on its inhibition. In previous studies, the two hypocholesterolemic peptides (LPYP and IAVPGEVA) were isolated and identified from soy protein. Based on these peptide sequences, a number of peptides were designed previously by using the correlation between the conformational flexibility and bioactivity. The design method that was applied in previous studies was slightly modified for the purpose of the current research and 5 tripeptides were designed and synthesized. Among the designed tripeptides, YIE showed the highest ability to inhibit HMGR. A kinetic analysis revealed that this peptide is a competitive inhibitor of HMG-CoA with an equilibrium constant of inhibitor binding (Ki) of 0.23 ± 0.03 μM. This is an overall 700-fold increase in inhibitory activity compared to the first isolated LPYP peptide from soybeans. The obtained correlation between the observed and predicted inhibitory activities is good enough in spite of the fact that a different length of peptides was used in the presented design. Statistical analysis suggests that the proposed approach is acceptable in the design of tripeptides.

Rational drug design of 6-substituted 4-anilino-2-phenylpyrimidines for exploration of novel ABCG2 binding site.

In the search for novel, highly potent, and nontoxic adjuvant chemotherapeutics to resolve the major issue of ABC transporter-mediated multidrug resistance (MDR), pyrimidines were discovered as a promising compound class of modern ABCG2 inhibitors. As ABCG2-mediated MDR is a major obstacle in leukemia, pancreatic carcinoma, and breast cancer chemotherapy, adjuvant chemotherapeutics are highly desired for future clinical oncology. Very recently, docking studies of one of the most potent reversers of ABCG2-mediated MDR were reported and revealed a putative second binding pocket of ABCG2. Based on this (sub)pocket, a series of 16 differently 6-substituted 4-anilino-2-phenylpyrimidines was designed and synthesized to explore the potential increase in inhibitory activity of these ABCG2 inhibitors. The compounds were assessed for their influence on the ABCG2-mediated pheophorbide A transport, as well as the ABCB1- and ABCC1-mediated transport of calcein AM. They were additionally evaluated in MDR reversal assays to determine their half-maximal reversal concentration (EC50). The 6-substitution did not only show increased toxicity against ABCG2-overexpressing cells in combination with SN-38 but also a negative influence on cell viability in general. Nevertheless, several candidates had EC50 values in the low double-digit nanomolar concentration range, qualifying them as some of the most potent reversers of ABCG2-mediated MDR. In addition, five novel multitarget ABCB1, ABCC1, and ABCG2 inhibitors were discovered, four of them exerting their inhibitory power against the three stated transporters at least in the single-digit micromolar concentration range.

Inhibitory activities of flavonoids from Eupatorium adenophorum against acetylcholinesterase

Fifteen flavonoids isolated from the Eupatorium adenophorum showed inhibitory activities against acetylcholinesterase (AChE) isolated from Caenorhabditis elegans and Spodoptera litura. Their IC50 values ranged from 12.54 to 89.06μg/mL and 12.08 to 86.01μg/mL, respectively against the AChE isolated from the nematode and insect species. AChE was inhibited in a dose-dependent manner by all tested flavonoids, The isolated compound quercetagetin-7-O-(6-O-caffeoyl-β-D-glucopyranoside) displayed the highest inhibitory effect against AChE from C. elegans and S. litura, with IC50 values of 12.54 μg/mL and 12.58 μg/mL, respectively. The structure-activity relationship of flavonoids on the inhibitory activities indicated that additional phenolic hydroxyl groups in the glucose were favorable for their inhibitory effects and the degree of increase in inhibitory activity also depended on the number of phenolic hydroxyl groups. The Lineweaver-Burk and Dixon plots indicated that quercetagetin-7-O-(6-O-caffeoyl-β-d-glucopyranoside) is a reversible inhibitor against AChE. Quercetagetin-7-O-(6-O-caffeoyl-β-d-glucopyranoside), 5,4′-Dihydroxytlavone and quercetin-3-O-β-d-glucopyranoside inhibited AChE in a mixed-type competitive manner and these compounds might be the dual binding site AChE inhibitors. Further, nine compounds showed poisonous effects against C. elegans and inhibitory effects on the growth and development of S. litura.

Comparative examination of levodopa pharmacokinetics during simultaneous administration with lactoferrin in healthy subjects and the relationship between lipids and COMT inhibitory activity in vitro

ABSTRACT Background: Lactoferrin (bLF) is an iron-binding multifunctional protein that is abundant in milk. In mice, it inhibits catechol-O-methyltransferase (COMT) activity and increases blood levodopa levels. However, the clinical effects are unknown. Objective: The objective of this study was to determine the effect of bLF on the kinetics of levodopa in blood. Design: The effects of the concomitant administration of a combined formulation of levodopa and an aromatic amino acid decarboxylase inhibitor and bLF on the concentration of levodopa in blood and its metabolism were assessed in eight healthy subjects. In addition, we analyzed the association with clinical factors and evaluated whether clinical factors affected the COMT inhibitory activity of bLF in vitro. Results: Although not statistically significant, the peak plasma concentration (C max) of levodopa increased by 18.5%. From the results of the stratified analysis of total cholesterol, a relationship with ΔC max was predicted. Therefore, bLF was reacted with cholesterol in the presence of lecithin and sodium deoxycholate in vitro to evaluate COMT inhibitory activity, and an increase in inhibitory activity was observed. By contrast, the ester compound cholesteryl oleate had no effect. The inhibitory activity of free fatty acids, which are known to interact with bLF, was also enhanced. Conclusion: The COMT inhibitory activity of bLF is not effective in elevating blood levodopa levels. However, in humans with high lipid levels, such as cholesterol, interactions may enhance the inhibitory effect, resulting in the enhanced absorption of levodopa. Trial registration: ID, UMIN000026787, registered 30 March 2017; URL, https://upload.umin.ac.jp/cgi-open-bin/ctr_e/ctr_view.cgi?recptno=R000030749 Trial registration: UMIN Japan identifier: UMIN000026787

Structure-Based Virtual Screening and Biological Evaluation of Peptide Inhibitors for Polo-Box Domain.

The polo-box domain of polo-like kinase 1 (PLK1-PBD) is proved to have crucial roles in cell proliferation. Designing PLK1-PBD inhibitors is challenging due to their poor cellular penetration. In this study, we applied a virtual screening workflow based on a combination of structure-based pharmacophore modeling with molecular docking screening techniques, so as to discover potent PLK1-PBD peptide inhibitors. The resulting 9 virtual screening peptides showed affinities for PLK1-PBD in a competitive binding assay. In particular, peptide 5 exhibited an approximately 100-fold increase in inhibitory activity (IC50 = 70 nM), as compared with the control poloboxtide. Moreover, cell cycle experiments indicated that peptide 5 effectively inhibited the expression of p-Cdc25C and cell cycle regulatory proteins by affecting the function of PLK1-PBD, thereby inducing mitotic arrest at the G2/M phase. Overall, peptide 5 can serve as a potent lead for further investigation as PLK1-PBD inhibitors.

Virtual screening identification and chemical optimization of substituted 2-arylbenzimidazoles as new non-zinc-binding MMP-2 inhibitors

Matrix metalloproteinases (MMPs) are a large family of zinc-dependent endoproteases known to exert multiple regulatory roles in tumor progression and invasiveness. This encouraged over the years the approach of MMP, and particularly MMP-2, targeting for anticancer treatment. Early generations of MMP inhibitors, based on aspecific zinc binding groups (ZBGs) assembled on (pseudo)peptide scaffolds, have been discontinued due to the clinical emergence of toxicity and further drawbacks, giving the way to inhibitors with alternative zinc-chelator moieties or not binding the catalytic zinc ion.In the present paper, we continue the search for new non-zinc binding MMP-2 inhibitors: exploiting previously identified compounds, a virtual screening (VS) campaign was carried out and led to the identification of a new class of ligands. The structure-activity relationship (SAR) of the benzimidazole scaffold was explored by synthesis of several analogues whose inhibition activity was tested with enzyme inhibition assays. By performing the molecular simplification approach, we disclosed different sets of single-digit micromolar inhibitors of MMP-2, with up to a ten-fold increase in inhibitory activity and ameliorated selectivity towards off-target MMP-8, compared to selected lead compound. Molecular dynamics calculations conducted on complexes of MMP-2 with docked privileged structures confirmed that analyzed inhibitors avoid targeting the zinc ion and dip inside the S1′ pocket. Present results provide a further enrichment of our insights for the design of novel MMP-2 selective inhibitors.

INHIBITION ACTIVITY OF BELIMBING WULUH(Averrhoa bilimbi linn) LEAF EXTRACT TO Streptococcus mutans ON ACRYLIC PLATE

Background : The leaf of Averrhoa bilimbi contains flavonoids and tannins as antibacterial agent against Streptococcus mutans . The extract of Averrhoa bilimbi leaf can be used as a natural alternative ingredients of denture cleanser. Purpose: The main purpose of this research is to know the different antibacterial activities of Averrhoa bilimbi leaf extract 6.25%, 12.5%, 25%, 50%, and 100% against Streptococcus mutans on acrylic plate. Methods: The method of this research was true experimental with posttest-only with control group design that consisted of Averrhoa bilimbi leaf extract in 6.25%, 12.5%, 25%, 50%, 100% and chlorhexidine gluconate 0.2%. The antibacterial testing method was using diffusion method. The measurement parameter was the amount of inhibition zone formed in the test medium. The data analysis was using Kruskall-Wallis test which resulted in p=0.000 (p<0.05). The result showed that there were differences in the inhibitory activity between each group. The test then continued with Mann Whitney test that results in p=0.009 (p<0.05) which showed meaningful differences between each group. Results: The smallest inhibitory zone of 6.25% was 10.08 mm, the biggest inhibitory zone of 100% was 23.07 mm, and the inhibitory zone of chlorhexidine gluconate in 0.2% was 25.05 mm. The lower extract concentration inhibitory zone were smaller than the high extract concentration. There was significant increase in inhibitory activity of Averrhoa bilimbi leaf extracts cause by the flavonoids and tannins content as antibacterial. Conclusion: It can be concluded that there are differences in the inhibitory activity of Averrhoa bilimbi leaf extract and chlorhexidine gluconate 0.2%.

Structure-based design, synthesis, and binding mode analysis of novel and potent chymase inhibitors

Based on insight from the X-ray crystal structure of human chymase in complex with compound 1, a lactam carbonyl of the diazepane core was exchanged with O-substituted oxyimino group, leading to amidoxime derivatives. This modification resulted in highly potent chymase inhibitors, such as O-phenylamidoxime 5f. X-ray crystal structure analysis indicated that compound 5f induced movement of the Leu99 and Tyr94 side chains at the S2 site, and the increase in inhibitory activity of O-phenyl amidoxime derivatives suggested that the O-phenyl moiety interacted with the Tyr94 residue. Surface plasmon resonance experiments showed that compound 5f had slower association and dissociation kinetics and the calculated residence time of compound 5f to human chymase was extended compared to that of amide compound 1.

In-sights into the effect of heavy metal stress on the endogenous mustard cystatin

Phytocystatins have been ascribed several protective roles against abiotic and biotic stress conditions. It was, therefore, thought worthwhile to document the effect of heavy metal stress on the endogenous plant cystatin. The mustard cystatin, purified from Brassica juncea (B. juncea) seeds retained its functional property of cysteine proteinase inhibition, despite exposure to high concentrations of metal ions, Cd2+ and Ni2+. An increase in inhibitory activity, ∼26% for Ni2+ and ∼16% for Cd2+ was observed, suggesting changes in protein conformation upon metal ion interaction. Isothermal calorimetric (ITC) studies show formation of a 1:1 binary complex on interaction with both metal ions but suggest a higher affinity for Ni2+. Fluorescence quenching data suggest a static quenching mechanism of interaction. Various spectroscopic analyses, namely, synchronous fluorescence, ANS fluorescence, far UV CD and ATR-FTIR spectroscopy show that the native mustard cystatin acquires a more ordered conformation upon interaction with metal ions. Differential Scanning Calorimetry indicates that the thermo-stability of the Ni2+ bound protein (Tm=109.4°C) is greater than both, the Cd2+ bound (Tm=104.5°C) and the native (Tm=99.5°C) forms. The B. juncea seed cystatin, is thus, identified as a potent and resilient member of the phytocystatin family with considerable inhibitory capacity despite exposure to heavy metal stress.

Synthesis, spectral characterization and inhibitory potency of ferrocene‐containing mannosides towards type 1 fimbriated Escherichia coli

Escherichia coli adhesion to the bladder epithelium is mediated through recognition of the tissue‐surface mannosylated proteins with bacterial lectin FimH. The inhibition of this recognition‐dependent interaction has been recognized as a promising strategy for the development of an anti‐adhesion therapy. Mannosides with either aryl‐ or elongated alkyl‐functionalized aglycon portions have been shown to be potent inhibitors of FimH‐mediated adhesion. Thus, we have synthesized four mannose‐based bioorganometallics containing an extended alkyl chain between sugar and ferrocene components connected via ester linkage (14 (n = 4) and 15 (n = 5)) or amide linkage (18 (n = 3) and 19 (n = 4)). The novel bioconjugates were characterized using infrared and NMR (1H, 13C, COSY, NOESY, HSQC, HMBC) spectroscopies, electrospray ionization mass spectrometry and high‐resolution mass spectrometry. Hemagglutination inhibitory assay of the novel bioorganometallics revealed an enhanced inhibitory potential in comparison to methyl α‐d‐mannoside. Thereby, the bioconjugate 19 exhibited a twofold increase in inhibitory activity compared with 14, 15 and 18. Copyright © 2016 John Wiley &amp; Sons, Ltd.

Identification of N-ethylmethylamine as a novel scaffold for inhibitors of soluble epoxide hydrolase by crystallographic fragment screening

Soluble epoxide hydrolase (sEH) is a potential target for the treatment of inflammation and hypertension. X-ray crystallographic fragment screening was used to identify fragment hits and their binding modes. Eight fragment hits were identified via soaking of sEH crystals with fragment cocktails, and the co-crystal structures of these hits were determined via individual soaking. Based on the binding mode, N-ethylmethylamine was identified as a promising scaffold that forms hydrogen bonds with the catalytic residues of sEH, Asp335, Tyr383, and Tyr466. Compounds containing this scaffold were selected from an in-house chemical library and assayed. Although the starting fragment had a weak inhibitory activity (IC50: 800μM), we identified potent inhibitors including 2-({[2-(adamantan-1-yl)ethyl]amino}methyl)phenol exhibiting the highest inhibitory activity (IC50: 0.51μM). This corresponded to a more than 1500-fold increase in inhibitory activity compared to the starting fragment. Co-crystal structures of the hit compounds demonstrate that the binding of N-ethylmethylamine to catalytic residues is similar to that of the starting fragment. We therefore consider crystallographic fragment screening to be appropriate for the identification of weak but promising fragment hits.

Synthetic Antimicrobial Peptides Exhibit Two Different Binding Mechanisms to the Lipopolysaccharides Isolated from Pseudomonas aeruginosa and Klebsiella pneumoniae.

Circular dichroism and 1H NMR were used to investigate the interactions of a series of synthetic antimicrobial peptides (AMPs) with lipopolysaccharides (LPS) isolated from Pseudomonas aeruginosa and Klebsiella pneumoniae. Previous CD studies with AMPs containing only three Tic-Oic dipeptide units do not exhibit helical characteristics upon interacting with small unilamellar vesicles (SUVs) consisting of LPS. Increasing the number of Tic-Oic dipeptide units to six resulted in five analogues with CD spectra that exhibited helical characteristics on binding to LPS SUVs. Spectroscopic and in vitro inhibitory data suggest that there are two possible helical conformations resulting from two different AMP-LPS binding mechanisms. Mechanism one involves a helical binding conformation where the AMP binds LPS very strongly and is not efficiently transported across the LPS bilayer resulting in the loss of inhibitory activity. Mechanism two involves a helical binding conformation where the AMP binds LPS very loosely and is efficiently transported across the LPS bilayer resulting in an increase in inhibitory activity. Mechanism three involves a nonhelical binding conformation where the AMP binds LPS very loosely and is efficiently transported across the LPS bilayer resulting in an increase in inhibitory activity.