Salicylic acid effect on the mechanism of Lelliottia amnigena causing potato soft rot

Salicylic acid (SA) plays an important role in protecting plants from biotic stresses. Lelliottia amnigena is a newly identified potato soft rot pathogen and there are no adequate studies on this soft rot pathogen. Therefore, this paper focussed on the effect of SA on the mechanism under which L. amnigena causes potato soft rot. L. amnigena was examined and detected to secrete pectinase, proteases, pectin lyase and cellulase, which are the most important pathogenic enzymes involved in the production of plant diseases. Sterilised healthy potato tubers were inoculated with 0.2 mL of L. amnigena suspension (3.69 CFU · mL−1 × 107 CFU · mL−1). After 24 h, 200 μL of four different SA concentrations (0.5 mM, 1.0 mM, 1.5 mM and 2.0 mM) were used to treat the tubers. Co-culture of L. amnigena and SA significantly reduced the activity of pectinase, protease, pectin lyase and cellulase by an average of 33.8%, 43.4%, 67.7% and 46.9%, across the four concentrations (0.5 mM, 1.0 mM, 1.5 mM and 2.0 mM), respectively, compared to the control. The average disease index was reduced by 54.7% across the four SA concentrations. Treatment with SA induced transcriptional levels of the superoxide dismutase, peroxide, catalase and glutathione S-transferase across the four levels by an average of 3.87, 3.25, 3.97 and 3.94-fold, respectively, compared to control. Based on our results, we could state that SA could reduce the activities of these extracellular enzymes produced by L. amnigena by modulating both enzymatic and non-enzymatic antioxidant activities and gene expression that induce natural resistance in plants against bacterial infections.

Purification of Pectin Lyase Enzyme from Bacillus pumilus Bacteria by Three-Phase Partitioning Method (TPP), Nanoflower Preparation and Investigation of Fruit Juice Clarification

In this study, firstly, Bacillus pumilus bacteria were isolated from tomato vegetables and identified. Then, the new pectin lyase enzyme was purified from B. pumilus, characterized, and hybrid nanoflower pectin lyase (hNF-PL) was synthesized the first time in this study. For this purpose, PL enzyme was produced in a solid culture medium using B. pumilus bacterium, and PL was purified in 191.8 folds with a yield of 78.2% using the three-phase partitioning (TPP) technique. Using SDS-PAGE, PL enzyme was determined to have a single subunit, and molecular weight was defined as 32.88 kDa with gel chromatography technique. This is the very first study to easily immobilize purified PL enzyme onto nanoflower chitosan/calcium pyrophosphate hybrid NPs. The synthesized nanoflower hNF-PL structure was characterized by SEM, FT-IR, XRD, and TEM chromatographic methods. In the final phase of the study, the effects of the pure PL and hNF-PL enzymes on the clarification and cleavage rate of fruit juices obtained from black grape, pomegranate, peach, red apple, and plum were investigated. Under the light shed by this study determined that the hNF-PL enzyme clarified the fruit juices more effectively than the pure PL enzyme.

An insight into the sequential changes in enzymatic activities during retting of jute (Corchorus spp. L.).

Aim: To study the dynamics of enzymes involved in biochemical process of jute (Corchorus spp.) retting with and without microbial retting consortium. Methodology: Two large scale retting trials were conducted with and without microbial retting consortium in triplicate. The retting water samples were collected every day at 24 hrs interval from both the trials. Polygalcturonase (PG), pectin lyase (PNL) and xylanase activities along with the pH were measured from the collected retting water samples following standard procedure. Fibre quality parameters were also studied from the resultant fibre obtained from both the retting trials. Results: There was a sharp decrease in pH of retting liquor by 1.35 units and that of pectin lyase activity by 97.9 Uml-1 within 24 hrs of inoculation of microbial retting consortium. Thereafter, higher pectin lyase (123.1 Uml-1), polygalacturonase (3.56 Iuml-1) and xylanase (0.818 IUml-1) activities were recorded during middle stage of retting. The enzyme activities were lower and non-significant at last stage of retting (11-14 days). The completion of retting without microbial consortium took longer time due to lower enzymatic activities as compared to microbial consortium mediated retting. Interpretation: The PG, PNL and xylanase enzymes released by the microbial consortium during retting of jute helped in faster biodegradation of pectin and xylan compared to control retting. Hence, the pre retting treatment of jute with microbial consortium is suggested for quick retting.

Effect of solid loading on the behaviour of pectin-degrading enzymes

Background Pectin plays a role in the recalcitrance of plant biomass by affecting the accessibility of other cell wall components to enzymatic degradation. Elimination of pectin consequently has a positive impact on the saccharification of pectin-rich biomass. This work thus focused on the behaviour of different pectin-degrading enzymes in the presence of low (5%) to high (35%) solid loading of lemon peel. Results High solid loading of lemon peel affected pectin solubilisation differently depending on the pectinase used. Pectin lyase was less sensitive to a reduction of water content than was a mixture of endopolygalacturonase and pectin methylesterase, regardless of whether or not the latter's mode of action is processive or not. Marked changes in water mobility were observed along with enzymatic degradation depending on the enzyme used. However, the pectin lyase resulted in less pronounced shifts in water distribution than polygalacturonase–pectin methylesterase mixtures. At similar pectin concentration, pectin solutions hindered the diffusion of hydrolases more than the solid substrate. This can be attributed to the high viscosity of the highly concentrated pectin solutions while the solid substrate may provide continuous diffusion paths through pores. Conclusions The increase in solid substrate loading reduced the efficiency of pectin-degrading enzymes catalysing hydrolysis more significantly than those catalysing β-elimination. LF-NMR experiments highlighted the impact of solid loading on water mobility. Compared to other enzymes and whatever the solid loading, pectin lyase led to longer relaxation times linked with the most destructuration of the solid substrate. This new information could benefit the biorefinery processing of pectin-rich plant material when enzymes are used in the treatment.

Pectin-Lyase-Modified Ginseng Extract and Ginsenoside Rd Inhibits High Glucose-Induced ROS Production in Mesangial Cells and Prevents Renal Dysfunction in db/db Mice

Diabetes increases the incidence rate of chronic renal disease. Pectin-lyase-modified ginseng (GS-E3D), with enhanced ginsenoside Rd content, has been newly developed. In this study, renal protective roles of GS-E3D in type-2 diabetic db/db mice were investigated. The generation of reactive oxygen species (ROS) induced by high glucose (25 mM) was reduced by ES-E3D (75%) and ginsenoside Rd (60%). Diabetic db/db mice received 100 or 250 mg/kg/day of GS-E3D daily via oral gavage for 6 weeks. Albuminuria and urinary 8-hydroxy-2′-deoxyguanosine (8-OhdG, an oxidative stress marker) levels were increased in db/db mice and the levels recovered after GS-E3D treatment. In renal tissues, TUNEL-positive cells were decreased after GS-E3D treatment, and the increased apoptosis-related protein expressions were restored after GS-E3D treatment. Therefore, GS-E3D has a potent protective role in diabetes-induced renal dysfunction through antioxidative and antiapoptotic activities. These results may help patients to select a dietary supplement for diabetes when experiencing renal dysfunction.

Synergism between Cutinase and Pectinase in the Hydrolysis of Cotton Fibers’ Cuticle

Scouring is one of the initial steps in the processing of natural textile fibers (e.g., cotton), performed to remove waxes and pectins, together with spinning oils and other impurities of the plant cell cuticle. Traditional chemical bleaching with boiling NaOH led to harsh removal of the entire fabric’s cuticle waxy layer accompanied by an unwanted alkaline waste. Extracellular lytic enzymes such as lipases, cellulases and pectinases play an essential role in host plant-pathogen interactions. They degrade the plant cuticle and tissue and enable pathogen invasion. Such enzymes, specifically cutinase and pectinase, have been considered potential bio-scouring agents to degrade the cotton fabric cuticle’s outer layer at low temperature and alleviate environmental pollution. In this work, the combined effect of cutinase, pectin lyase, or polygalacturonase on the scouring of cotton fabrics was studied using evaporative light-scattering reverse-phase HPLC and GC-MS analysis of the reaction components, and measuring changes in the cotton fabrics’ properties. The traditional method of cotton fabrics’ scouring with NaOH resulted in decreased pectin content and increased cellulose fibers accessibility, evaluated by specific staining. Treating the cotton fibers’ cuticle with cutinase led to the acidification of the reaction mixture, a decrease in enzyme-specific activity, and elevation in hexadecanoic acid and octadecanoic acids in the reaction fluid. These two saturated fatty acids are the main wax constituents of raw cotton fabrics, identified using GC-MS after dichloromethane reflux overnight. Treating cotton fabrics with each of the three enzymes, cutinase, pectin lyase, or polygalacturonase, increased their pectin removal, as measured by high concentrations of D-galacturonic acid and other pectin constituents in the reaction fluid. A synergistic effect was found in the combined treatment of cutinase and pectin lyase in the hydrolysis of the cotton fibers’ cuticle. This effect was expressed in high water absorbency of the treated fibers, increased fabric weight loss and sharp elevation of a cutin and pectin monomer’s related peaks (retention time [RT] = 4.1 min and 2.9, 4.5 min, respectively). A model was suggested for the synergistic action between cutinase and pectin lyase. It assumes that the cuticle’s digestion by cutinase results in the enlargement and formation of outer layer micropores, which enables the rapid penetration of pectinase into the inner pectin layer.

Immobilization of Purified Pectin Lyase from Acinetobacter calcoaceticus onto Magnetic Carboxymethyl Cellulose Nanoparticles and Its Usability in Food Industry

An important component of the pectinase enzyme complex is pectin lyase (polymethylgalacturonate lyase; EC 4.2.2.10). In this study, extracellular pectin lyase enzyme was produced from Acinetobacter calcoaceticus bacteria. Pectin lyase was then purified using three-phase precipitation (TPP) technique with 25.5% yield. The pectin lyase was immobilized covalently via the L-glutaraldehyde spacer to the carboxymethyl cellulose. The immobilized pectin lyase was magnetized using Fe3O4 nanoparticles. Purified pectin lyase was connected to magnetized support material after 90 min at the rate of 80%. The most appropriate immobilization conditions were determined as pH 8 and 30°C. By characterizing the free and immobilized enzyme, KM, Vmax, and optimum pH and optimum temperature values were determined. It was optimum pH 8 and temperature 50°C for both free and immobilized pectin lyase. The structural characterization of the immobilized pectin lyase modified with Fe3O4 nanoparticles was carried out by SEM, FT-IR, and XRD chromatographic analyses. At the end of the study, free and immobilized enzymes were used for purification of some fruit juices and results were compared.