Effects of simulated acid rain on the physiology of carmine spider mite, Tetranychus cinnabarinus (Boisduvals) (Acari: Tetranychidae)

Abstract:  The physiological effect of simulated acid rain sprayed on carmine spider mite Tetranychus cinnabarinus (Boisduvals) and host plant, were measured in a series of laboratory trials. We examined potential changes in three kinds of protective enzymes [peroxidase (POD), superoxide dismutase (SOD) and catalase (CAT)] and three hydrolases [acid phosphatase (ACP), alkaline phosphatase (ALP) and carboxylesterase (CarE)] in response to changes in pH values of simulated acid rain at different time of exposure. POD, SOD and CAT activities increased significantly with the increase in the acidity of the acid rain, reaching the highest levels at pH 4.0 or 3.0, and then declined. Changes in ACP activity were similar to those observed in the protective enzymes. The increasing extent of the activities of these four enzymes after 30 and 45 days treatment became smaller than that after 15 days treatment . ALP activities decreased as pH value declined. There were no significant changes in CarE activities after 15 and 45 days, but that in pH 4.0 and 3.0 decreased after 30 days. The enhanced anti-oxidation enzyme levels (POD, SOD and CAT) and ACP activities in pH 4.0 and 3.0 reduced the effects of these toxic products on mites, resulting in the strengthening of the defensive power, and increase in survival and reproductive power of the mites, thus leading to an increase in the density of mites on host plant. From these results, we inferred that POD, SOD, CAT and ACP might be relevant to population changes of mites under acid rain pressure.     

Characterization of surface binding sites in glycoside hydrolases: A computational study

Structural properties of carbohydrate surface binding sites (SBSs) were investigated with computational methods. Eighty‐five SBSs of 44 enzymes in 119 Protein Data Bank (PDB) files were collected as a dataset. On the basis of SBSs shape, they were divided into 3 categories: flat surfaces, clefts, and cavities (types A, B, and C, respectively). Ligand varieties showed the correlation between shape of SBSs and ligands size. To reduce cut‐off differences in each SBSs with different ligand size, molecular docking were performed. Molecular docking results were used to refine SBSs classification and binding sites cut‐off. Docking results predicted putative ligands positions and displayed dependence of the ligands binding mode to the structural type of SBSs. Physicochemical properties of SBSs were calculated for all docking results with YASARA Structure. The results showed that all SBSs are hydrophilic, while their charges could vary and depended to ligand size and defined cut‐off. Surface binding sites type B had highest average values of solvent accessible surface area. Analysis of interactions showed that hydrophobic interactions occur more than hydrogen bonds, which is related to the presence of aromatic residues and carbohydrates interactions.     

A New Enzymatic Reaction: Enzyme Catalyzed Ammonolysis of Carboxylic Esters

A new enzymatic reaction of carboxylic esters and ammonia (ammonolysis) was studied. This reaction provides a synthetically useful and mild alternative for the synthesis of amides. Several lipases and one esterase acted as catalyst. Ammonolysis of esters of chiral carboxylic acids gave higher ee values than hydrolysis under comparable reaction conditions. Furthermore, consecutive enzymatic esterification and ammonolysis provided a convenient one-pot synthesis of carboxylic amides from carboxylic acids.     

Lipase‐catalyzed enantioselective transesterification of prochiral 1‐((1,3‐dihydroxypropan‐2‐yloxy)methyl)‐5,6,7,8‐tetrahydroquinazoline‐2,4(1H,3H)‐dione in ionic liquids

The application of ionic liquids as solvents for transesterification of prochiral pirymidine acyclonucleoside using lipase (EC 3.1.1.3) Amano PS from Burkholderia cepacia (BCL) is reported. The effect of using medium reaction, acyl group donor, and temperature on the activity and enantioselectivity of BCL was studied. From the investigated ionic solvents, the hydrophobic ionic liquid [BMIM]PF₆] was the preferred medium for enzymatic reactions. However, the best result was obtained in the mixture [BMIM][PF₆]:TBME (1:1 v/v) at 50°C. Enzyme activity and selectivity in [BMIM][PF₆]:TBME (1:1 v/v) was slightly higher in than in conventional organic solvents (for example, TBME), and in this condition, good activity and enantioselectivity were associated with unique properties of ionic liquid such as hydrophobicity and high polarity. Independently of solvents, monester of (R)‐configuration was obtained in excess. Under optimal conditions, desymmetrization of the prochiral compound using different acyl donors was performed. If vinyl butyrate was used as the acylating agent, BCL completely selectively acylated enantiotopic hydroxyl groups.     

Characterization of chitinases and β-1,3-glucanases in grapefruit flavedo during fruit development

Chitinase (EC 3.2.1.14) and β-1,3-glucanase (EC 3.2.1.39) activities in the flavedo of grapefruit ( Citrus paradisi cv. Marsh) were determined at 17 times during the course of fruit development. Chitinase activity is initially high in flavedo, but drops rapidly and is low, although fairly constant throughout the remainder of fruit development. In contrast to chitinase, β-1,3-glucanase activity is lowest in young fruit and increases during development. Western blots of crude flavedo extracts following SDS-PAGE were probed with antibodies raised against purified citrus chitinase and β-1,3-glucanase. Results of immunostaining revealed that changes in the activities of chitinase and β-1,3-glucanase were reflected in the amount of chitinase and glucanase protein present in the extracts. Only a single chitinase band was detected on western blots of crude flavedo extracts, whereas one glucanase band was present in young fruit and a second one appeared later in older fruit. Partial purification of flavedo chitinases and glucanases was performed using extracts prepared from immature and mature fruit for the two enzymes, respectively. Acidic and basic forms of both enzymes were present in the extracts; acidic and basic forms of chitinase were present in nearly equal amounts whereas basic glucanases predominated (91% of total activity). Acidic and basic chitinases differed in substrate specificity as well as products of degradation indicating the heterogeneous nature of the enzymes. Both acidic and basic glucanases required the presence of β-1,3 linkages for activity, were active against both soluble and insoluble β-1,3 glucans and generated similar products.     

Biochemical and Immunological Relationships Between Endo-β-1,4-Glucanases from Cockroaches

The cellulase from Geoscapheus dilatatus consisted of two major and four minor endo-β-1,4-glucanase components. Two major and one minor component were purified to homogeneity. The major endo-β-1,4-glucanase components, named GD1 and GD2, were similar to EG1 and EG2 from Panesthia cribrata in terms of M_r and kinetic properties. The purified minor component, named GD3, was distinct from GD1 and GD2 because of a lower M_r and a lower specific activity. Polyclonal antibodies raised against the two major endo-β-1,4-glucanase components, EG1 and EG2, of the cellulase from P. cribrata cross-reacted with each other and with pure GD1 and GD2 from the foregut and midgut of the related cockroach G. dilatatus but did not cross-react with GD3. Endo-β-1,4-glucanase components were partially purified from the foregut and midgut of four other cockroaches. These comprised three other Australian cockroaches also from the superfamily Blaberoidea and one American cockroach, Cryptocercus punctulatus, from the superfamily Blattoidea. The endogenous cellulases from all cockroaches examined consisted of either two or three major endo-β-1,4-glucanase components. The amino acid sequence of the N-terminus region of the two major endo-β-1,4-glucanase components from P. cribrata were determined and are homologous with those belonging to glycosyl hydrolase family 9 (cellulase family E).     

Alpha/beta‐hydrolases: A unique structural motif coordinates catalytic acid residue in 40 protein fold families

The alpha/beta‐hydrolases are a family of acid‐base‐nucleophile catalytic triad enzymes with a common fold, but using a wide variety of substrates, having different pH optima, catalyzing unique catalytic reactions and often showing improved chemical and thermo stability. The ABH enzymes are prime targets for protein engineering. Here, we have classified active sites from 51 representative members of 40 structural ABH fold families into eight distinct conserved geometries. We demonstrate the occurrence of a common structural motif, the catalytic acid zone, at the catalytic triad acid turn. We show that binding of an external ligand does not change the structure of the catalytic acid zone and both the ligand‐free and ligand‐bound forms of the protein belong to the same catalytic acid zone subgroup. We also show that the catalytic acid zone coordinates the position of the catalytic histidine loop directly above its plane, and consequently, fixes the catalytic histidine in a proper position near the catalytic acid. Finally, we demonstrate that the catalytic acid zone plays a key role in multi‐subunit complex formation in ABH enzymes, and is involved in interactions with other proteins. As a result, we speculate that each of the catalytic triad residues has its own supporting structural scaffold, similar to the catalytic acid zone, described above, which together form the extended catalytic triad motif. Each scaffold coordinates the function of its respective catalytic residue, and can even compensate for the loss of protein function, if the catalytic amino acid is mutated.