• Journal of Microbiology and Biotechnology
• /
• v.16 no.8
• /
• pp.1192-1200
• /
• 2006

An apple polygalacturonase-inhibiting protein (PGIP), which specifically inhibits endopolygalacturonase (PG, EC 3.2.1.15) from Botryosphaeria dothidea, was purified from Botryosphaeria dothidea-infected apple (Malus domestica cv. Fuji) fruits. The purified apple PGIP had a molecular mass of 40 kDa. The N-terminal amino acid sequence of the purified protein showed high homologies to those of PGIP from pear (100%), tomato (70%), and bean (65%). We also purified polygalacturonase (PG) from B. dothidea. The PG hydrolyzes pectic components of plant cell walls. When the extracted apple pectic cell wall material was treated with purified apple PGIP and B. dothidea PG, the amount of uronic acid released was lower than that treated with B. dothidea PG alone. This result demonstrates that PGIP functions specifically by inhibiting cell wall maceration of B. dothidea PG Furthermore, we characterized the de novo function of the PGIP against PG on the solubilization and depolymerization of polyuronides from cell wall of apple fruits inoculated with B. dothidea. This result demonstrated that the PGIP of plants exhibits one of the direct defense mechanisms against pathogen attack by inhibiting PGs that are released from pathogens for hydrolysis of cell wall components of plants.

• Korean Journal Plant Pathology
• /
• v.11 no.4
• /
• pp.312-317
• /
• 1995

The polygalacturonase (PG) production in rotten apples by Botryosphaeria dothidea was purified by using gel filtration and ion exchange column chromatography, and the biochemical characters of PG were investigated. The purified PG appeared as a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) with approximate molecular weight of 49 kilodalton (kDa). The molecular weight was equal to the native molecular weight estimated by gel filtration. The Km and Vmax values of PG were 0.51 mg/ml and 90.9 $\mu$M/min/ml, respectively. Optimum pH was 4.0~5.0, and the PG activity was stable from pH 5.0~10.0. Optimum temperature of the enzyme activity was 4$0^{\circ}C$. The PG activity was relatively stable at 2$0^{\circ}C$, but it was reduced 45% at 4$0^{\circ}C$ and completely inactivated at 8$0^{\circ}C$. The PG activity was considerably inhibited by Cu2+, Zn2+, SDS and EDTA, whereas it was not effected by Ca2+, K+, Mg2+ or Na+ ions.

• Korean Journal of Plant Tissue Culture
• /
• v.25 no.2
• /
• pp.131-134
• /
• 1998

$\textrm{T}_{5}$ progeny of one transgenic tomato line (To9) carrying antisense polygalacturonase (PG) cDNA was generated by selfing. Five $\textrm{T}_{5}$ plants were used to analyse in detail. The PG antisense gene was stably inherited through fifth generations. In all five $\textrm{T}_{5}$ plants, expression of the antisense transcripts were detected. In consequence, it led to a reduction of the PG enzyme activity in ripe fruit to between 37% and 65% that of normal. In two plants the expression of endogenous PG gene was inhibited in ripe fruit.

• Korean Journal Plant Pathology
• /
• v.10 no.3
• /
• pp.215-221
• /
• 1994

Polygalacturonase (PG) produced by Botrytis cinerea in the culture broth containing citrus pectin as a carbon source was partially purified and characterized. PG was produced on a range of carbon sources such as starch, glycerol, cellobiose, and Na+-PAG with total activities of 34.8, 32.0, 29.2, 27.8 units, respectively. The specific activity was highest with 2316.7 units on Na+-PGA. Proteins of culture filtrate were concentrated with polyethylene glycol and acetone and applied to a hydroxyapatite column. Among three active fractions collected from the column, the reaction containing the highest PG activity was resolved by a Q-sepharose column. The active fraction from the Q-sepharose column was further purified by HPLC Mono Q column. The partially purified enzyme was analyzed by 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Among a few protein bands revealed, the amount of the protein of which molecular weight estimated to be 43 kDa coincided with the PG activity. The partially purified PG had optimal temperatures between 35~55$^{\circ}C$ and pH between 4.5~5.5.

• Journal of Sensor Science and Technology
• /
• v.30 no.5
• /
• pp.337-341
• /
• 2021

In this paper, we describe the fabrication of a paper-based enzyme-linked immunosorbent assay (ELISA) to detect polygalacturonase (PG), which is used as a biomarker to determine whether a plant is infected with a disease. The proposed paper-based ELISA can analyze the concentration of PG in a short time using a small sample compared to the traditional ELISA, which is generally performed using a well plate. To increase the resolution of the sensor, we optimized the dilution ratio of the HRP-conjugated goat anti-rabbit IgG antibody and the dilution ratio of the anti-PG and HRP-conjugated goat anti-rabbit IgG antibodies. Furthermore, for quantitative analysis of PG concentration, Delta RGB analysis was conducted to detect color changes in the sensing window displayed by the PG samples at various concentrations. Based on the experiment, the fabricated paper-based ELISA could measure at least 0.25 ㎍ of PG and the measurement range was 0.25-2 ㎍. Therefore, the paper-based ELISA for detecting PG is expected to be able to determine the presence or absence of disease in crops at the infection stage in the future.

• Korean Journal of Plant Tissue Culture
• /
• v.22 no.6
• /
• pp.351-355
• /
• 1995

A truncated Polygalacturonase (PG) cDNA was fused in reverse orientation to the CaMV 35S promoter of the binary vector pCA643, and introduced into tomato cells by Agrobaderium - mediated transformation. Transformed cells were selected using kanamycin as select agent then regenerated into plants. After selfed, one transgenic line (T9), was germinated and grown on MS medium containing 1 mg/mL of kanamycin Genomic Southern analysis of a T9 progeny with labelled PG2 cDNA probe showed a single antisense PC fragment as well as the endogenous PG2 gene, suggesting that PC antisense gene was integrated into tomato genome. Northern blot analysis demonstrated that the antisense RNA was produced from the transgene at much tiger level than the endogenous PG2 gene. Polygalacturonase activity analysis of the fruit from transgenic plants demonstrated that the antisense transgene expression caused 4 to 60% reduction of endogenous PG activity.

• Korean Journal of Food Science and Technology
• /
• v.25 no.5
• /
• pp.576-581
• /
• 1993

Polygalacturonase(PG) was extracted from Chinese cabbage and partially purified by ammonium sulfate fractionation and ion-exchange chromatography. Three fractions, D-PG, C-1 PG, and C-2 PG, were separated by CM-Sephadex C-25 column chromatography and FPLC Mono Q HR 5/5 or Mono S HR 5/5 column and their physicochemical characteristics were investigated. All three fractions had optimum pH and temperature of 4.5 and $65^{\circ}C$, and were stable in the range of pH $4.5{\sim}8.0$. These fractions were inhibited by 0.8mM $CaCl_2$ or 0.6M NaCl. In the thermal inactivation of PC isozymes at $65{\sim}80^{\circ}C$, z-values were $8.4{\sim}9.3^{\circ}C$ and D-values at $80^{\circ}C$ were In the range of $120{\sim}126s$. Thermodynamic constants were calculated from thermal inactivation data of the isozymes and were applicable to estimation of thermal process time of retort pouched Kimchi.

• Journal of Life Science
• /
• v.16 no.4
• /
• pp.653-658
• /
• 2006

An apple polygalacturonase-inhibiting protein (PGIP), that specifically inhibited endopolygalacturonase (PG, EC 3.2.1.15) from Botryosphaeria dothidea, was purified from B. dothidea infected apple (Malus domestica cv. Fuji) fruits. The apple PGIP was a mixed-type inhibitor of PG from B. dothidea. Optimal temperature for the maximum enzyme activity was $40^{\circ}C$, and optimum pH of the purified PGIP was pH 5.0. PGIP was stable up to temperature of $60^{\circ}C$ and was completely suppressed after heating at $70^{\circ}C$ for 10 min, PGIP was stable at pH between 4 and 8. Inhibition of PG by PGIP was reduced by $K^+$, $Cu^{2+}$, $Mg^{2+}$, $Ca^{2+}$ and $Zn^{2+}$ metal ion, sodium dodecyl sulfate (SDS) and 1,2-diaminocyclohexane tetra acetate (CDTA).

• The Korean Journal of Food And Nutrition
• /
• v.9 no.1
• /
• pp.52-58
• /
• 1996

This study was carried out 19 investigate the characteristics of pectinesterase (PE), polygalacturonase (PG), lipoxygenase(LOX) and peroxidase (POD) in hot pepper to know the effect of hot pepper on food quality during food processing and storage. The results were as follows : 1. The optimum pH of PE was pH 7.5 and the activity of PE below pH 5.5 was revealed scarcely, The concentration of NaCl and $CaCl_2$ that showed the highest activity of PE were 0.2M and 0.05M, respectively. 2. The optimum pH of PG was pH 6.0 and the activity of PG in acidity was higher than that in alkalinity. The activity of PG was maximum at 0.3M NaCl and 0.2mM $CaCl_2$. Above the concentration of 0.5M NaCl and 0.5M $CaCl_2$, the activity of PG was lower than that of PG not adding these salts 3. The optimum pH of LOX was pH 7.0 and pH 8.5. 4. The optimum pH of POD was pH 6.0 and the activity of POD was higher in weak acidity and neutrality than in alkalinity. POD activity was slightly decreased by the increase of NaCl and $CaCl_2$ concentration.

• Korean Journal of Food Science and Technology
• /
• v.16 no.1
• /
• pp.41-46
• /
• 1984

The pectic enzymes produced from Aspergillus niger were separated into three fractions (F-A, F-I and F-II) by means of Sephadex and DEAF-Sephadex column chromatography. Each enzyme fraction was characterized by determining viscosity change and reducing surgar of the pectic acid-enzyme mixture and analyzing thin layer chromatogram of the reaction products. F-I rapidly reduced the viscosity of pectic acid solution and released reducing groups in a random manner so that appeared to be an endo-polygalacturonase (endo-PG). The optimum pH of endo-PG for viscosity reducing activity was 4.2 and that for releasing reducing surgar was 4.7. In the thermal inactivation of endo-PG of $30-45^{\circ}C$, the enthalpy of activation was 217.3 kj/mole and z-value was $7.5^{\circ}C$. F-II and F-A were determined as endo-polymethylgalacturonase and exo-polygalacturonase, respectively.