Circulating proalbumin associated with a variant proteinase inhibitor

The unique finding of normal proalbumin in human plasma provides an insight into the mechanism of propeptide cleavage. Proalbumin, present as 1–5% of the total albumin, was found in a boy whose prime problem was the presence of a mutant proteinase inhibitor, α₁-antitrypsin Pittsburgh (358Met→Arg) [2]. The infeerred structure of human proalbumin was confirmed as ArgGlyValPheArgArgAlb. On incubation with various enzymes (trypsin, tryptase, thrombin, chymotrypsin, chymase and cathepsin B), only trypsin was capable of converting proalbumin to albumin. There was no conversion when proalbumin was incubated with whole blood, plasma or serum. However, intravenous injection of proalbumin into a rat resulted in complete conversion to albumin, the half-life of this process being 6 h. We conclude that propeptide cleavage is dependent on a serine proteinase which is inhibited intracellularly, by the mutant inhibitor, and that all the albumin in the boy was secreted as proalbumin, but was subjected to a separate cleavage process after export from the hepatocyte.     

MicroRNA-506 inhibits the proliferation and invasion of mantle cell lymphoma cells by targeting B7H3

Background

Aberrant expression of B7 homologue 3 (B7H3) has been observed in various malignancies. Our previous study demonstrated that knocking down of B7H3 inhibited cell proliferation, invasion and enhanced the therapeutic efficacy of chemotherapy in mantle cell lymphoma (MCL). However, the mechanism regulating of B7H3 expression remains unknown. Here, we present a new regulatory microRNA of B7H3, miR-506, that directly targets B7H3 and may play an inhibitory role in MCL progression.

C-terminus of heat shock protein 60 can activate macrophages by lectin-like oxidized low-density lipoprotein receptor 1

Immune responses against antigens generally require an efficient activation of antigen-presenting cells (APCs). Currently, the targeting of vaccine antigens to APCs has emerged as a promising strategy for boosting vaccine immunogenicity. Here, we reported that the C-terminus of heat shock protein 60 (HSP60C) can activate mouse peritoneal macrophages to secret a series of cytokines, and phosphorylation of p38 mitogen-activated protein kinase (MAPK) and NF-κB p65 was involved in the pathway. We showed that the activation effect of HSP60C on macrophages was independent of toll-like receptor (TLR) 4 and the TLR-associated myeloide differentiation factor 88 (MyD88). Knockdown of lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) reduced the activation of HSP60C-induced macrophage p38 MAPK, NF-κB p65 and cytokine secretion to some extent. Finally, we found that HSP60C up-regulated the expression of LOX-1 on macrophages and ovalbumin (OVA) model antigen fused with HSP60C markedly enhanced OVA-specific IgG responses. Thus, our results unravel a novel LOX-1-dependent pathway by which HSP60C can effectively activate macrophages and APCs targeting based on LOX-1 interaction is a promising approach to improve vaccines.     

Intramolecular C(sp3)H amination of arylsulfonyl azides with engineered and artificial myoglobin-based catalysts

The direct conversion of aliphatic CH bonds into CN bonds provides an attractive approach to the introduction of nitrogen-containing functionalities in organic molecules. Following the recent discovery that cytochrome P450 enzymes can catalyze the cyclization of arylsulfonyl azide compounds via an intramolecular C(sp³)H amination reaction, we have explored here the CH amination reactivity of other hemoproteins. Various heme-containing proteins, and in particular myoglobin and horseradish peroxidase, were found to be capable of catalyzing this transformation. Based on this finding, a series of engineered and artificial myoglobin variants containing active site mutations and non-native Mn- and Co-protoporphyrin IX cofactors, respectively, were prepared to investigate the effect of these structural changes on the catalytic activity and selectivity of these catalysts. Our studies showed that metallo-substituted myoglobins constitute viable CH amination catalysts, revealing a distinctive reactivity trend as compared to synthetic metalloporphyrin counterparts. On the other hand, amino acid substitutions at the level of the heme pocket were found to be beneficial toward improving the stereo- and enantioselectivity of these Mb-catalyzed reactions. Mechanistic studies involving kinetic isotope effect experiments indicate that CH bond cleavage is implicated in the rate-limiting step of myoglobin-catalyzed amination of arylsulfonyl azides. Altogether, these studies indicate that myoglobin constitutes a promising scaffold for the design and development of CH amination catalysts.     

Investigations on unconventional hydrogen bonds in RNA binding proteins: The role of CH⋯OC interactions

We have investigated the roles played by CH⋯OC interactions in RNA binding proteins. There was an average of 78 CH⋯OC interactions per protein and also there was an average of one significant CH⋯OC interaction for every 6 residues in the 59 RNA binding proteins studied. Main chain–Main chain (MM) CH⋯OC interactions are the predominant type of interactions in RNA binding proteins. The donor atom contribution to CH⋯OC interactions was mainly from aliphatic residues. The acceptor atom contribution for MM CH⋯OC interactions was mainly from Val, Phe, Leu, Ile, Arg and Ala. The secondary structure preference analysis of CH⋯OC interacting residues showed that, Arg, Gln, Glu and Tyr preferred to be in helix, while Ala, Asp, Cys, Gly, Ile, Leu, Lys, Met, Phe, Trp and Val preferred to be in strand conformation. Most of the CH⋯OC interacting polar amino acid residues were solvent exposed while, majority of the CH⋯OC interacting non polar residues were excluded from the solvent. Long and medium-range CH⋯OC interactions are the predominant type of interactions in RNA binding proteins. More than 50% of CH⋯OC interacting residues had a higher conservation score. Significant percentage of CH⋯OC interacting residues had one or more stabilization centers. Sixty-six percent of the theoretically predicted stabilizing residues were also involved in CH⋯OC interactions and hence these residues may also contribute additional stability to RNA binding proteins.     

Precursor De13.1 from Conus delessertii defines the novel G gene superfamily

Peptide de13a was previously purified from the venom of the worm-hunting cone snail Conus delessertii from the Yucatán Channel, México. This peptide has eight cysteine (Cys) residues in the unique arrangement CCCCCCCC, which defines the cysteine framework XIII (“” represents one or more non-Cys residues). Remarkably, δ-hydroxy-lysine residues have been found only in conotoxin de13a, which also contains an unusually high proportion of hydroxylated amino acid residues. Here, we report the cDNA cloning of the complete precursor De13.1 of a related peptide, de13b, which has the same Cys framework and inter-Cys spacings as peptide de13a, and shares high protein/nucleic acid sequence identity (87%/90%) with de13a, suggesting that both peptides belong to the same conotoxin gene superfamily. Analysis of the signal peptide of precursor De13.1 reveals that this precursor belongs to a novel conotoxin gene superfamily that we chose to name gene superfamily G. Thus far superfamily G only includes two peptides, each of which contains the same, distinctive Cys framework and a high proportion of amino acid residues with hydroxylated side chains.     

Unlocking nature’s CH bonds

In an idealistic setting, it can be imagined that if every CH bond on an organic molecule could be selectively functionalized, the fields of chemical synthesis and drug discovery would be forever revolutionized. With the purpose of investigating the practicality of this idealistic scenario, our group has endeavored to unlock the potential of nature’s CH bonds by developing palladium-catalyzed, site selective CH insertions that can be incorporated into both known and new catalytic cycles. To this end, we have developed a number of catalytic transformations that not only provide rapid diversification of simple starting materials and natural products through CH functionalization, but streamline the synthesis of a variety of natural products with biological activity and expand upon methods to access highly valuable enantiopure materials.     

Development of indazole mineralocorticoid receptor antagonists and investigation into their selective late-stage functionalization

The derivatization of pharmaceuticals is a core activity in the discovery and development of new medicines. Late-stage functionalization via modern CH functionalization chemistry has emerged as a powerful technique with which to diversify advanced pharmaceutical intermediates. We report herein a case study in late-stage functionalization towards the development of a new class of indazole-based mineralocorticoid receptor antagonists (MRA). An effort to modify the electronics of the core indazole heterocycle inspired the use of modern CH borylation chemistry. New reactivity patterns were revealed and studied computationally. Ultimately, a de novo synthesis delivered a key 6-fluoroindazole compound 26, a potent MRA with excellent metabolic stability.     

Metal heterodiene complexes

In this article a survey is given of novel results in the field of the chemistry of α-diimine complexes of some metal atoms of the main group, transition metal and post transition metal area. It is shown that α-diimines of the type RNC(R′)(R″)CNR (= R-DAB) and 2-C₅H₄NC(R′)NR (= R-Pyca) are very versatile with regards to both their coordination chemistry and chemical reactivity when coordinated to a metal or metal cluster. The type of reaction occurring depends critically on the type of metal(s), the other ligands present on the metal(s), the type of coordination of the α-diimines, and particularly on the steric and electronic properties of R,R′ and R″ situated on the R-DAB and R-Pyca ligands.     

Facile synthesis of 2-aryl 5-hydroxy benzo[d]oxazoles and their in vitro anti-proliferative effects on various cancer cell lines

2-Aryl 5-hydroxy benzo[d]oxazoles were designed as potential anticancer agents. A one-pot synthesis of these compounds dispenses the need for ortho-disubstituted precursor, aminophenol and proceeds via CN formation as a key step followed by CO cyclization to form benzo[d]oxazoles. The single crystal X-ray diffraction study was used to confirm the molecular structure of a representative compound unambiguously. All of these compounds were evaluated for their anti-proliferative properties in vitro against five cancer cell lines as well as noncancerous cells. Most of these compounds showed selective growth inhibition of cancer cells and few of them were found to be promising with IC₅₀ values in the range of 0.8–2.8?μM, comparable to the known anticancer drug doxorubicin.     

FTIR, 1H NMR and EPR spectroscopy studies on the interaction of flavone apigenin with dipalmitoylphosphatidylcholine liposomes

Apigenin (5,7,4′-trihydroxyflavone) is a cancer chemopreventive agent and a member of the family of plant flavonoids. Apigenin interaction with liposomes formed with dipalmitoylphosphatidylcholine (DPPC) was investigated by means of FTIR spectroscopy, ¹H NMR and EPR techniques. Fluorescent microscopy and electron microscopy were applied to study the apigenin effects on colon myofibroblasts and human skin fibroblasts. The strong rigidifying effect of apigenin with respect to polar head groups was concluded on the basis of the action of the flavone on partition coefficient of Tempo spin label between the water and lipid phases. The ordering effect was also found in hydrophobic region at the depth monitored by 5-SASL and 16-SASL spin labels. The inclusion of apigenin to the membrane restricted the motional freedom of polar head groups lowering penetration of Pr^3 + ions to the membranes. The ¹H NMR technique supported also the restriction of motional freedom of the membrane in the hydrophobic region, especially in the zone of CH₂ groups of alkyl chains. FTIR analysis showed that apigenin incorporates into DPPC liposomes via hydrogen bonding between its own hydroxyl groups and lipid polar head groups in the COPOC segment. It is also very likely that hydroxyl groups of apigenin link with polar groups of DPPC by water bridges. Electron and fluorescence microscopic observations revealed changes in the internal membrane organization of the examined cells. In conclusion, the changes of the structural and dynamic properties of membranes can be crucial for processes involving tumor suppression signal transduction pathways and cell cycle regulation.     

Water-soluble N-carboxymethylchitosan derivatives: Preparation,characteristics and its application

In this work, only N-substituted chitosan derivatives (water-soluble N-carboxymethylchitosan derivatives: N-CMC) with different degrees of substitution were obtained by reaction of a fully deacetylated chitosan (derived from deacetylation of chitosan using decrystallized method) with monochloroacetic acid at pH 8 and temperature of 90 °C. The structure of N-carboxymethylchitosan and chitosan was characterized by IR, ¹H, ¹³C and ¹H–¹³C NMR-HSQC spectra. In the IR spectrum of the N-carboxymethylchitosan, the appearance of peak at 1742 cm^?1 was assigned for CO group of NHCH₂COOH of substituted chitosan. In the ¹H NMR spectra, the peaks at about 3.81÷4.06 ppm, assigned for CH₂ groups of NHCH₂ and N(CH₂)₂, were the major feature, while in the ¹H–¹³C NMR-HSQC spectra, signals of CH₂ confirmed the presence of these two different substituted CH₂ groups. The degree of substitution (DS) of N-monosubstitution (DS_N-mono) decreased from 0.47 to 0.03 meanwhile that of N,N-disubstitution (DS_N,N-di) increased from 0.52 to 0.96 since the mass ratio of chitosan/monochloroacetic acid changing from 1/1 to 1/4. The N-carboxymethylchitosan derivatives have been used for adsorption Cu(II) ion from aqueous solution. The results shown that the optimum conditions for adsorption Cu(II) ion in nitrate solution were pH 6.5, temperature of 30 °C, for 60–90 min and the substituted chitosan derivative having DS_N-mono of 0.16 and DS_N,N-di of 0.81 had maximum adsorption capacity of 192 mg Cu(II) per gram of N-CMC.     

Crystal and molecular conformation of the fluorescent probe. 8-anilino-1-naphthalenesulfonic acid (ANS).

The crystal and molecular structure of the fluorescent probe 8-anilino-1-naphthalenesulfonic acid (ANS) has been determined as the ammonium monohydrate with two conformationally distinct molecules in the triclinic $P\overline{1}$ lattice. The angles between the aromatic rings and the CNC plane are ?9/22° and 112/22° respectively. There is an NH...O intramolecular hydrogen bond in each molecule indicating that hydrogen bond formation is not dependent on the anilino geometry. There are also short intramolecular H...H contacts involving the hydrogens which have anomalous proton shifts shown in a recent NMR study.     

Vancomycin resistance: Modeling backbone variants with d-Ala-d-Ala and d-Ala-d-Lac peptides

To seek vancomycin analogs with broader antibacterial activity, effects of backbone modifications for the agylcon 2 on binding with d-Ala-d-Ala- and d-Ala-d-Lac-containing peptides were investigated by Monte Carlo/free energy perturbation (MC/FEP) calculations. The experimental trend in binding affinities for 2 with three tripeptides was well reproduced. Possible modifications of the peptide bond between residues 4 and 5 were then considered, specifically for conversion of the OCNH linkage to CH₂NH₂⁺ (6), FCCH (7), HCCH (8), and HNCO (9). The MC/FEP results did not yield binding improvements for 7, 8, and 9, though the fluorovinyl replacement is relatively benign. The previously reported analog 6 remains as the only variant that exhibits improved affinity for the d-Ala-d-Lac sequence and acceptable affinity for the d-Ala-d-Ala sequence.     

Development of a safe and scalable route towards a tau PET tracer precursor

A scalable 5-step synthesis of the diazacarbazole derivative 1 used as tau PET tracer precursor is reported. Key features of this synthesis include a Buchwald-Hartwig amination, a Pd catalyzed CH activation and a Suzuki-Miyaura cross-coupling.     

Synthesis and characterization of cis-dioxomolybdenum(VI) complexes with sterically bulky tripodal tetradentate ligands

A new series of sterically bulky tripodal tetradentate ligands have been synthesized. The ligands were found to react readily with MoO ₂(acac) ₂ in absolute methanol to yield the corresponding cis- dioxoMo(VI) complexes. These complexes were characterized by IR, ¹H and ¹³C NMR, UV-Vis spectroscopy, elemental analysis and cyclic voltammetry (CV). Cyclic voltammetry was performed in DMF and CH ₂Cl ₂. In CH ₂Cl ₂ the ease of reduction was found to decrease in the order MoO ₂[LNO ₂ (SCH ₃)]> MoO ₂[LNO ₂(OCH ₃)]> MoO ₂ [LNO ₂ (N(CH ₃) ₂)]. The CV results are consistent with that expected for a S,O,N substitution pattern.     

Geometrical effects of phospholipid olefinic bonds on the structure and dynamics of membranes: A molecular dynamics study

The trans isomers of fatty acids are found in human adipose tissue. These isomers have been linked with deleterious health effects (e.g., coronary artery disease). In this study, we performed molecular dynamics simulations to investigate the structures and dynamic properties of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylcholine (POPC) and 1-palmitoyl-2-elaidoyl sn-glycero-3-phosphatidylcholine (PEPC) lipid bilayers. The geometry of the olefinic bond and membrane packing effects significantly influenced the conformations and dynamics of the two CC single bonds adjacent to the olefinic bond. For the PEPC lipid, the two CC single bonds adjacent to the olefinic bond adopted mainly nonplanar skew–trans and planar cis–trans motifs; although the cis conformation featured relatively strong steric repulsion, it was stabilized through membrane packing because its planar structure is more suitable for membrane packing. Moreover, membrane packing effects stabilized the planar transition state for conformational conversion to a greater extent than they did with the nonplanar transition state, thereby affecting the dynamics of conformational conversion. The rotational motions of the first neighboring CC single bonds were much faster than those of typical saturated CC single bonds; in contrast, the rotational motions of the second neighboring CC single bonds were significantly slower than those of typical saturated torsion angles. The packing of PEPC lipids is superior to that of POPC lipids, leading to a smaller area per lipid, a higher order parameter and a smaller diffusion coefficient. The distinct properties of POPC and PEPC lipids result in PEPC lipids forming microdomains within a POPC matrix.     

Characterizations of divalent lanthanoid iodides in tetrahydrofuran by UV-Vis,fluorescence and ESR spectroscopy

Spectrochemical properties of LnI ₂ (Ln = Sm, Eu, Yb) prepared with lanthanoid metal and diiodoethane in THF were characterized by means of UV—Vis, fluorescence and ESR spectroscopy. A comparative investigation has also been made concerning LnI ₃ (Ln= LaLu) and the Grignard-type compounds RLnI (R = Et, Ph; Ln = Eu, Yb). LnI ₂ (Ln = Sm, Eu, Yb) was also prepared from reaction of lanthanoid metal and iodine in THF.