Alpha(1)-acid glycoprotein: an acute phase protein with inflammatory and immunomodulating properties

alpha(1)-Acid glycoprotein (AGP) is a protein with a molecular weight of 41-43 kDa and is heavily glycosylated (45%). Due to the presence of sialic acids, it is negatively charged (pI=2.7-3.2). AGP is an acute phase protein in all mammals investigated to date. The serum concentration of AGP rises several fold during an acute phase response, the systemic answer to a local inflammatory stimulus. Also, its glycosylation pattern can change depending on the type of inflammation. The biological function of this protein is not clear. A number of activities on different type of blood cells have been described. In vivo, AGP clearly has protective effects in several models of inflammation. Here we review the data supporting an anti-inflammatory and immunomodulating role of AGP.     

Discovery, synthesis, and structure-activity studies of tetrazole based growth hormone secretagogues

A novel class of Growth Hormone Secretagogues (GHS), based on a tetrazole template, has been discovered. In vitro SAR and in vivo potency within this new class of GHS are described. The tetrazole 9q exhibits good oral bioavailability in rats and dogs as well as efficacy following an oral 10 mg/kg dose in dogs. Solution and solid phase protocols for the synthesis of tetrazole based GHS have been developed.     

Design and synthesis of novel pyrimidone analogues as HIV-1 integrase inhibitors

A series of novel pyrimidone analogues have been designed and synthesized as HIV-1 integrase (IN) inhibitors. This study demonstrated that introducing a substituent in the N1-position of the pyrimidone scaffold does not significantly influence IN inhibitory activity. Molecular docking studies showed these compounds could occupy the IN active site and form pi–pi interactions with viral DNA nucleotides DC16 and DA17 to displace reactive viral DNA 3′OH and block intasome activity.     

Ligand Photo-Isomerization Triggers Conformational Changes in iGluR2 Ligand Binding Domain

Neurological glutamate receptors bind a variety of artificial ligands, both agonistic and antagonistic, in addition to glutamate. Studying their small molecule binding properties increases our understanding of the central nervous system and a variety of associated pathologies. The large, oligomeric multidomain membrane protein contains a large and flexible ligand binding domains which undergoes large conformational changes upon binding different ligands. A recent application of glutamate receptors is their activation or inhibition via photo-switchable ligands, making them key systems in the emerging field of optochemical genetics. In this work, we present a theoretical study on the binding mode and complex stability of a novel photo-switchable ligand, ATA-3, which reversibly binds to glutamate receptors ligand binding domains (LBDs). We propose two possible binding modes for this ligand based on flexible ligand docking calculations and show one of them to be analogues to the binding mode of a similar ligand, 2-BnTetAMPA. In long MD simulations, it was observed that transitions between both binding poses involve breaking and reforming the T686-E402 protein hydrogen bond. Simulating the ligand photo-isomerization process shows that the two possible configurations of the ligand azo-group have markedly different complex stabilities and equilibrium binding modes. A strong but slow protein response is observed after ligand configuration changes. This provides a microscopic foundation for the observed difference in ligand activity upon light-switching.     

Identification of a Chemoreceptor for Tricarboxylic Acid Cycle Intermediates: DIFFERENTIAL CHEMOTACTIC RESPONSE TOWARDS RECEPTOR LIGANDS*

We report the identification of McpS as the specific chemoreceptor for 6 tricarboxylic acid (TCA) cycle intermediates and butyrate in Pseudomonas putida. The analysis of the bacterial mutant deficient in mcpS and complementation assays demonstrate that McpS is the only chemoreceptor of TCA cycle intermediates in the strain under study. TCA cycle intermediates are abundantly present in root exudates, and taxis toward these compounds is proposed to facilitate the access to carbon sources. McpS has an unusually large ligand-binding domain (LBD) that is un-annotated in InterPro and is predicted to contain 6 helices. The ligand profile of McpS was determined by isothermal titration calorimetry of purified recombinant LBD (McpS-LBD). McpS recognizes TCA cycle intermediates but does not bind very close structural homologues and derivatives like maleate, aspartate, or tricarballylate. This implies that functional similarity of ligands, such as being part of the same pathway, and not structural similarity is the primary element, which has driven the evolution of receptor specificity. The magnitude of chemotactic responses toward these 7 chemoattractants, as determined by qualitative and quantitative chemotaxis assays, differed largely. Ligands that cause a strong chemotactic response (malate, succinate, and fumarate) were found by differential scanning calorimetry to increase significantly the midpoint of protein unfolding (T_m) and unfolding enthalpy (ΔH) of McpS-LBD. Equilibrium sedimentation studies show that malate, the chemoattractant that causes the strongest chemotactic response, stabilizes the dimeric state of McpS-LBD. In this respect clear parallels exist to the Tar receptor and other eukaryotic receptors, which are discussed.     

α-Amylase from germinating soybean (Glycine max) seeds – Purification,characterization and sequential similarity of conserved and catalytic amino acid residues

Starch hydrolyzing amylase from germinated soybeans seeds (Glycine max) has been purified 400-fold to electrophoretic homogeneity with a final specific activity of 384 units/mg. SDS–PAGE of the final preparation revealed a single protein band of 100 kDa, whereas molecular mass was determined to be 84 kDa by MALDI–TOF and gel filtration on Superdex-200 (FPLC). The enzyme exhibited maximum activity at pH 5.5 and a pI value of 4.85. The energy of activation was determined to be 6.09 kcal/mol in the temperature range 25–85 °C. Apparent Michaelis constant (K_m(app)) for starch was 0.71 mg/mL and turnover number (k_cat) was 280 s^?1 in 50 mM sodium acetate buffer, pH 5.5. Thermal inactivation studies at 85 °C showed first-order kinetics with rate constant (k) equal to 0.0063 min^?1. Soybean α-amylase showed high specificity for its primary substrate starch. High similarity of soybean α-amylase with known amylases suggests that this α-amylase belongs to glycosyl hydrolase family 13. Cereal α-amylases have gained importance due to their compatibility for biotechnological applications. Wide availability and easy purification protocol make soybean as an attractive alternative for plant α-amylase. Soybean can be used as commercially viable source of α-amylase for various industrial applications.