Dynamics of the protein structure of T169S pyranose 2‐oxidase in solution: Molecular dynamics simulation

Pyranose 2‐oxidase (P2O) from Trametes multicolor contains FAD as cofactor, and forms a tetramer. The protein structure of a mutated P2O, T169S (Thr169 is replaced by Ser), in solution was studied by means of molecular dynamics simulation and analyses of photoinduced electron transfer (ET) from Trp168 to excited isoalloxazine (Iso*), and was compared with wild type (WT) P2O. Hydrogen bonding between Iso and nearby amino acids was very similar as between T169S and WT protein. Distances between Iso and Tyr456 were extremely heterogeneous among the subunits, 1.7 (1.5 in WT) in subunit A (Sub A), 0.97 (2.2 in WT) in Sub B, 1.3 (2.1 in WT) in Sub C, 1.3 nm (2.0 in WT) in Sub D. Mean values of root of mean square fluctuation over all residues were greater by four times than those in WT. This suggests that the protein structure of T169S is much more flexible than that of WT. Electrostatic (ES) energies between Iso anion in one subunit and ionic groups in the entire protein were evaluated. It was found that more than 50% of the total ES energy in each subunit is contributed from other subunits. Reported fluorescence decays were analyzed by a method as WT, previously reported. Electron affinities of Iso* in T169S were appreciably higher than those in WT. Static dielectric constants near Iso and Trp168 were also quite higher in T169S than those in WT.     

Entrapment of 5‐aminolevulinic acid under edible composite film of konjac glucomannan and chitosan

5‐Aminolevulinic acid (5‐ALA) is a known plant regulator and growth promoter. It is a very sensitive and highly unstable compound that is easy to deteriorate. Here we propose a novel approach to stabilize 5‐ALA into a film. Films from konjac glucomannan (KGM), KGM treated with alkali solution (KGOH), chitosan (CHI) as well as blends between KGOH and CHI were fabricated for 5‐ALA entrapment. It was found that the efficiency of KGM film, KGOH film and CHI film for 5‐ALA entrapment was 55.7 ± 0.73%, 58.3 ± 0.36% and 60.3 ± 0.18 %, respectively. A 25:75 (%w/w) blended film (KGOH/CHI) showed the highest entrapment efficiency of 5‐ALA (65.9 ± 0.37%) versus other films. The possible mechanism for entrapment of 5‐ALA in blended film was postulated under two mechanisms. A secondary amide that leads to the interaction between the amino group of CHI and carboxyl group of 5‐ALA is proposed as the first mechanism. The fact that the 5‐ALA molecule was entrapped within the complexity of KGOH structure is proposed as the second mechanism. Therefore, stabilizing 5‐ALA in a film may be an alternative way to use and preserve 5‐ALA for further applications.     

Role of the electrostatic energy between the photo-products and ionic groups on the photoinduced electron transfer rates from aromatic amino acids to the excited flavin in five single-point substitution isoforms of the charged amino acid residue-13 in the FMN-binding protein

Effect of the charge (negative, positive or neutral) of amino acid residue-13 on the photoinduced electron transfer (ET) from Trp32, Tyr35 and Trp106 to the excited isoalloxazine was evaluated in the flavin mononucleotide-binding protein from Desulfovibrio vulgaris isolate Miyazaki F (DvFBP). The protein structures of the wild type and the four isoforms where glutamic acid-13 is replaced with lysine (E13K), arginine (E13R), threonine (E13T) and glutamine (E13Q) in aqueous solution were obtained by molecular dynamics simulation. The distances between the amino acid residue-13 and isoalloxazine (Iso), and between the amino acid residue-13 and the ET donors were longer than 1 nm. The ET rates were evaluated with the Kakitani and Mataga model (KM theory) from their ultrafast fluorescence dynamics by means of a non-linear least squares method. Electrostatic (ES) energies between the photo-products and other ionic groups in the proteins markedly varied among ET donors and among the DvFBP isoforms, while the other physical quantities related to the ET rates, the solvent reorganisation and ES energies between the Iso anion and the donor cations did not vary much between the proteins and donors. A plot of the logarithmic ET rates versus either the total free energy gaps or the net ES energies between the photo-products and the other ionic groups both displayed a parabolic function and so the net ES energies are an important influential factor upon the ET rate, in addition to the donor–acceptor distance.     

Binding mode prediction of biologically active compounds from plant Salvia Miltiorrhiza as integrase inhibitor

Integrase (IN), an essential enzyme for HIV-1 replication, has been targeted in antiretroviral drug therapy. The emergence of HIV-1 variants clinically resistant to antiretroviral agents has lead to the development of alternative IN inhibitors. In the present work, binding modes of a high potent IN inhibitor, M₅22 and M₅32, within the catalytic binding site of wild type (WT) IN were determined using molecular docking calculation. Both M₅22 and M₅32 displayed similar modes of binding within the IN putative binding pocket and exhibited favorable interactions with the catalytic Mg²⁺ ions, the nearby amino acids and viral DNA through metal-ligand chelation, hydrogen bonding and π-π stacking interactions. Furthermore, the modes of action of these two compounds against the mutated Y212R, N224H and S217H PFV IN were also predicted. Although the replacement of amino acid could somehow disturb inhibitor binding mode, almost key interactions which detected in the WT complexes were fairly conserved. Detailed information could highlight the application of M₅22 and M₅32 as candidate IN inhibitors for drug development against drug resistant strains.     

Melissopalynological analysis of stingless bee (Tetragonula pagdeni) honey in Eastern Thailand

Melissopalynological analysis of 72 Tetragonula pagdeni honey samples, collected from various locations in Chanthaburi (A) and Trat (B) provinces, Eastern Thailand during March 2015, was performed. Overall, 45 pollen types species belonging to 22 plant families were identified. The predominant pollen type was from Nephelium lappaceum and comprised 48.5% of the pollen in honey from location A2 (Pathavee district) and 45.3% in location B1 (AoYai district). The secondary pollen types, Wodyetia bifurcata and Mimosa pudica, accounted for 20.1% and 17.0%, respectively, in location B3 (Nhongsamed district), while Cocos nucifera accounted for 17.2% in location A2. In addition, pollen types of C. nucifera, M. pudica, N. lappaceum, Asystasia gangetica, Amaranthus lividus, Areca catechu, Chromolaena odorata and Durio zibethinus were found in T. pagdeni honey from all sampled locations. Furthermore, in the dearth period, T. pagdeni foraged food (as in pollen was present) from Musa sp., Acacia mangium and various weed species, such as A. gangetica, A. lividus, Ageratum conyzoides, Bidens pilosa, C. odorata, Melampodium divaricatum, Mikania cordata, Merremia umbellata, M. pudica, Pennisetum pedicellarum and Thysanoleana maxima, from within a 500?m radius around the hive to maintain their colonies.