Activity cliffs were systematically extracted from public domain X-ray structures of targets for which complexes with multiple ligands were available, following the concept of three-dimensional (3D) cliffs. Binding modes of ligands with well-defined potency measurements were compared in a pairwise manner, and their 3D similarity was calculated using a previously reported property density function-based method taking conformational, positional, and chemical differences into account. Requiring the presence of at least 80% 3D similarity and a potency difference of at least 2 orders of magnitude as cliff criteria, a total of 216 well-defined 3D activity cliffs were detected in the Protein Data Bank (PDB). These 3D-cliffs involved a total of 269 ligands active against 38 different targets belonging to 17 protein families. For 255 of these compounds, binding modes were available at high crystallographic resolution. All 3D-cliffs were analyzed in detail and assigned to different categories on the basis of crystallographic interaction patterns. In many instances, differences in ligand-target interactions suggested plausible causes for origins of 3D-cliffs. In other cases, short-range interactions seen in X-ray structures were insufficient to deduce possible reasons for cliff formation. The 3D-cliffs described herein further advance the rationalization of activity cliffs at the level of ligand-target interactions and should also be useful for other applications such as the calibration of energy functions for structure-based design. The pool of identified activity cliffs is provided to enable subsequent structure-based analyses of cliffs.

Publicly available kinase inhibitors have been analyzed in detail. Nearly 19000 inhibitors have been identified with activity against 266 different kinases. Thus, about half of the human kinome is currently covered with active small molecules. The distribution of inhibitors across the kinome is uneven. Most available kinase inhibitors are likely to be type I inhibitors. By contrast, type II inhibitors are rare but usually have high potency. Kinase inhibitors generally display high scaffold diversity. Activity cliffs with an at least 100-fold difference in potency are only found for inhibitors of 106 kinases, which is partly due to only small numbers of compounds available for many kinases, in addition to scaffold diversity. Moreover, kinase inhibitors are less promiscuous than often thought. More than 70% of available inhibitors are only annotated with a single kinase activity, and only ∼1% of the inhibitors are active against five or more kinases.

Using the example of cathepsin K, we demonstrate the design of highly potent and selective azadipeptide nitrile inhibitors. A systematic scan with respect to P2 and P3 substituents was carried out. Structural modifications strongly affected the enzyme-inhibitor association (but not dissociation) rate. A combination of optimized P2 and P3 substituents with a methylation of the P3-P2 amide linker resulted in the picomolar cathepsin K inhibitor 19 with remarkable selectivity over cathepsins L, B, and S.

A coumarin-tetrahydroquinoline hydride 8 was synthesized as a chemical tool for fluorescent labeling. The rigidified tricyclic coumarin structure was chosen for its suitable fluorescence properties. The connection of 8 with a vinyl sulfone building block was accomplished by convergent synthesis thereby leading to the coumarin-based, tripeptidomimetic activity-based probe 10, containing a Gly-Phe-Gly motif. Probe 10 was evaluated as inactivator of the therapeutically relevant human cysteine cathepsins S, L, K, and B: it showed particularly strong inactivation of cathepsin S. The detection of recombinant and native cathepsin S was demonstrated by applying 10 to in-gel fluorescence imaging.

No abstract

The enzyme catalyzing the ring-contracting conversion of the flavanonol taxifolin to the auronol alphitonin in the course of flavonoid degradation by the human intestinal anaerobe Eubacterium ramulus was purified and characterized. It stereospecifically catalyzed the isomerization of (؉)-taxifolin but not that of (؊)-taxifolin. The K m for (؉)-taxifolin was 6.4 ؎ 0.8 M, and the V max was 108 ؎ 4 mol min ؊1 (mg protein) ؊1 . The enzyme also isomerized (؉)-dihydrokaempferol, another flavanonol, to maesopsin. Inspection of the encoding gene revealed its complete identity to that of the gene encoding chalcone isomerase (CHI) from E. ramulus. Based on the reported X-ray crystal structure of CHI (M. Gall et al., Angew Chem Int Ed 53:1439 -1442, 2014, http://dx.doi.org/10.1002/anie.201306952), docking experiments suggest the substrate binding mode of flavanonols and their stereospecific conversion. Mutation of the active-site histidine (His33) to alanine led to a complete loss of flavanonol isomerization by CHI, which indicates that His33 is also essential for this activity. His33 is proposed to mediate the stereospecific abstraction of a proton from the hydroxymethylene carbon of the flavanonol C-ring followed by ring opening and recyclization. A flavanonol-isomerizing enzyme was also identified in the flavonoid-converting bacterium Flavonifractor plautii based on its 50% sequence identity to the CHI from E. ramulus. IMPORTANCE Chalcone isomerase was known to be involved in flavone/flavanone conversion by the human intestinal bacterium E. ramulus.Here we demonstrate that this enzyme moreover catalyzes a key step in the breakdown of flavonols/flavanonols. Thus, a single isomerase plays a dual role in the bacterial conversion of dietary bioactive flavonoids. The identification of a corresponding enzyme in the human intestinal bacterium F. plautii suggests a more widespread occurrence of this isomerase in flavonoid-degrading bacteria. F lavonoids represent a major group of plant-derived polyphenolic compounds and have been implicated in beneficial effects on human health (1-4). The flavonoids are classified into several subgroups, which include, among others, flavonols, flavanonols, flavones, flavanones, and chalcones. Quercetin (Fig. 1) is a highly abundant dietary flavonoid that shows a broad range of biological activities. Thus, this flavonol has been intensively studied with respect to its role in disease prevention and use in therapy (5-7). The effects of quercetin and other flavonoids depend on how they are metabolized in the human body, including their conversion by intestinal bacteria (8). Beside deglycosylation and deconjugation, intestinal bacteria are also able to further transform the resulting flavonoid aglycones. However, knowledge about the corresponding bacterial species and, in particular, the enzymes involved is still limited.Eubacterium ramulus and Flavonifractor plautii (formerly Clostridium orbiscindens) are human intestinal bacteria that have been demonstrated to cleave the central heterocyc...

No abstract

No abstract