The role of CH/π interactions in the high affinity binding of streptavidin and biotin.

The streptavidin-biotin complex has an extraordinarily high affinity (Ka: 1015mol-1) and contains one of the strongest non-covalent interactions known. This strong interaction is widely used in biological tools, including for affinity tags, detection, and immobilization of proteins. Although hydrogen bond networks and hydrophobic interactions have been proposed to explain this high affinity, the reasons for it remain poorly understood. Inspired by the deceased affinity of biotin observed for point mutations of streptavidin at tryptophan residues, we hypothesized that a CH/π interaction may also contribute to the strong interaction between streptavidin and biotin. CH/π interactions were explored and analyzed at the biotin-binding site and at the interface of the subunits by the fragment molecular orbital method (FMO) and extended applications: PIEDA and FMO4. The results show that CH/π interactions are involved in the high affinity for biotin at the binding site of streptavidin. We further suggest that the involvement of CH/π interactions at the subunit interfaces and an extended CH/π network play more critical roles in determining the high affinity, rather than involvement at the binding site.

Application of the fragment molecular orbital method analysis to fragment-based drug discovery of BET (bromodomain and extra-terminal proteins) inhibitors.

The molecular interactions of inhibitors of bromodomains (BRDs) were investigated. BRDs are protein interaction modules that recognizing ε-N-acetyl-lysine (εAc-Lys) motifs found in histone tails and are promising protein-protein interaction (PPI) targets. First, we analyzed a peptide ligand containing εAc-Lys to evaluate native PPIs. We then analyzed tetrahydroquinazoline-6-yl-benzensulfonamide derivatives found by fragment-based drug design (FBDD) and examined their interactions with the protein compared with the peptide ligand in terms of the inter-fragment interaction energy. In addition, we analyzed benzodiazepine derivatives that are high-affinity ligands for BRDs and examined differences in the CH/π interactions of the amino acid residues. We further surveyed changes in the charges of the amino acid residues among individual ligands, performed pair interaction energy decomposition analysis and estimated the water profile within the ligand binding site. Thus, useful insights for drug design were provided. Through these analyses and considerations, we show that the FMO method is a useful drug design tool to evaluate the process of FBDD and to explore PPI inhibitors.

Synthesis and structure-activity relationships of 1-benzylindane derivatives as selective agonists for estrogen receptor beta.

The estrogen receptor beta (ERß) selective agonist is considered a promising candidate for the treatment of estrogen deficiency symptoms in ERß-expressing tissues, without the risk of breast cancer, and multiple classes of compounds have been reported as ERß selective agonists. Among them, 6-6 bicyclic ring-containing structures (e.g., isoflavone phytoestrogens) are regarded as one of the cyclized analogues of isobutestrol 5b, and suggest that other cyclized scaffolds comprising 5-6 bicyclic rings could also act as selective ERß ligands. In this study, we evaluated the selective ERß agonistic activity of 1-(4-hydroxybenzyl)indan-5-ol 7a and studied structure-activity relationship (SAR) of its derivatives. Some functional groups improved the properties of 7a; introduction of a nitrile group on the indane-1-position resulted in higher selectivity for ERß (12a), and further substitution with a fluoro or a methyl group to the pendant phenyl ring was also preferable (12b, d, and e). Subsequent chiral resolution of 12a identified that R-12a has a superior profile over S-12a. This is comparable to diarylpropionitrile (DPN) 5c, one of the promising selective ERß agonists and indicates that this indane-based scaffold has the potential to provide better ERß agonistic probes.

Ab initio studies on the structure of and atomic interactions in cellulose III(I) crystals.

The crystal structure of cellulose III(I)was analyzed using first-principles density functional theory (DFT). The geometry was optimized using variable-cell relaxation, as implemented in Quantum ESPRESSO. The Perdew-Burke-Ernzerhof (PBE) functional with a correction term for long-range van der Waals interactions (PBE-D) reproduced the experimental structure well. By using the optimized crystal structure, the interactions existed among the cellulose chains in the crystal were precisely investigated using the NBO analysis. The results showed that the weak bonding nature of CH/O and the hydrogen bonding occur among glucose molecules in the optimized crystal structure. To investigate the strength of interaction, dimeric and trimeric glucose units were extracted from the crystal, and analyzed using MP2 ab initio counterpoise methods with BSSE correction. The results estimated the strength of the interactions. That is, the packed chains along with a-axis interacts with weak bonding nature of CH/O and dispersion interactions by -7.50 kcal/mol, and two hydrogen bonds of O2HO2…O6 and O6HO6…O2 connect the neighboring packed chains with -11.9 kcal/mol. Moreover, FMO4 calculation was also applied to the optimized crystal structure to estimate the strength of the interactions. These methods can well estimate the interactions existed in the crystal structure of cellulose III(I).

Structure-activity relationship studies of 5-benzylaminoimidazo[1,2-c]pyrimidine-8-carboxamide derivatives as potent, highly selective ZAP-70 kinase inhibitors.

Zeta-associated protein, 70 kDa (ZAP-70), a spleen tyrosine kinase (Syk) family kinase, is normally expressed on T cells and natural killer cells and plays a crucial role in activation of the T cell immunoresponse. Thus, selective ZAP-70 inhibitors might be useful not only for treating autoimmune diseases, but also for suppressing organ transplant rejection. In our recent study on the synthesis of Syk family kinase inhibitors, we discovered that novel imidazo[1,2-c]pyrimidine-8-carboxamide derivatives possessed potent ZAP-70 inhibitory activity with good selectivity for ZAP-70 over other kinases. In particular, compound 26 showed excellent ZAP-70 kinase inhibition and high selectivity for ZAP-70 over structurally related Syk. The discovery of a potent, highly selective ZAP-70 inhibitor would contribute a new therapeutic tool for autoimmune diseases and organ transplant medication.

The importance of CH/pi hydrogen bonds in rational drug design: An ab initio fragment molecular orbital study to leukocyte-specific protein tyrosine (LCK) kinase.

The interaction energy was calculated, by the ab initio FMO method, for complexes between LCK protein and four inhibitors (staurosporine, BMS compound 2, and our compounds 3 and 4). In every case a number of CH/pi hydrogen bonds have been disclosed in the so-called adenine pocket. In complexes of 2, 3, and 4, CH/pi and NH/pi hydrogen bonds have been observed in another pocket. In view of the above results, the aniline ring of 3 was replaced by 2,6-dimethyl aniline to increase the potency for LCK kinase. A 10-fold increase in the potency has been achieved for 4 over 3. We suggest that the concept of weak hydrogen bonds is useful in the rational design of drugs.

Structure-activity relationship studies of imidazo[1,2-c]pyrimidine derivatives as potent and orally effective Syk family kinases inhibitors.

Spleen tyrosine kinase (Syk) and zeta-associated protein kinase of 70k Da (ZAP-70) are members of the Syk family and non-receptor-type protein tyrosine kinases, which play crucial roles in B- and T-cell activation. Therefore, a Syk family tyrosine kinases inhibitor would be a useful therapeutic agent for the treatment of various allergic disorders and autoimmune diseases. Previously, we reported that 1,2,4-triazolo[4,3-c]pyrimidine derivative 1 and 1,2,4-triazolo[1,5-c]pyrimidine derivative 2 showed strong inhibitory activities against Syk family kinases. These compounds also exhibited high-level suppression of IL-2 in cellular assays. However, their oral efficacies were poor in a mouse model of IL-2 production. To improve oral effectiveness, we investigated a new series of Syk family kinases inhibitors. We found that imidazo[1,2-c]pyrimidine derivatives potently inhibited the Syk family kinases. Among these agents, compound 9f not only showed strong inhibitory activities against Syk and ZAP-70 kinases in vitro, but its oral administration resulted in the in vivo suppression of both the passive cutaneous anaphylaxis reaction and Concanavalin A-induced IL-2 production in a mouse model.

A novel Syk family kinase inhibitor: design, synthesis, and structure-activity relationship of 1,2,4-triazolo[4,3-c]pyrimidine and 1,2,4-triazolo[1,5-c]pyrimidine derivatives.

Splenic tyrosine kinase (Syk) family kinases, which are members of the protein tyrosine kinase family, play crucial roles in immune responses, with Syk participating in B-cell activation and the zeta-associated protein 70 kDa (ZAP-70) kinase being involved in T-cell activation. Therefore, Syk family kinase inhibitors are candidate therapeutic agents for the treatment of various allergic disorders and autoimmune diseases. We designed 1,2,4-triazolo[4,3-c]pyrimidine and 1,2,4-triazolo[1,5-c]pyrimidine derivatives as Syk family kinase inhibitors, based on literature reports and structure-based drug design. These derivatives showed significant Syk inhibitory activities, with ZAP-70 inhibition. Representative compounds 10d and 11 not only exhibited strong inhibition of both Syk and ZAP-70 kinase but also suppressed IL-2 production by peripheral blood mononuclear cells and whole blood.