Two Extracellular Poly(ε-caprolactone)-Degrading Enzymes From Pseudomonas hydrolytica sp. DSWY01T: Purification, Characterization, and Gene Analysis
Poly(ε-caprolactone) (PCL) is an artificial polyester with commercially promising application. In this study, two novel PCL-degrading enzymes named PCLase I and PCLase II were purified to homogeneity from the culture supernatant of an effective polyester-degrading bacterium, Pseudomonas hydrolytica...
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Frontiers Media S.A.
2022-03-01
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| Series: | Frontiers in Bioengineering and Biotechnology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fbioe.2022.835847/full |
| _version_ | 1819002191618244608 |
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| author | Linying Li Linying Li Xiumei Lin Jianfeng Bao Hongmei Xia Hongmei Xia Fan Li Fan Li |
| author_facet | Linying Li Linying Li Xiumei Lin Jianfeng Bao Hongmei Xia Hongmei Xia Fan Li Fan Li |
| author_sort | Linying Li |
| collection | DOAJ |
| description | Poly(ε-caprolactone) (PCL) is an artificial polyester with commercially promising application. In this study, two novel PCL-degrading enzymes named PCLase I and PCLase II were purified to homogeneity from the culture supernatant of an effective polyester-degrading bacterium, Pseudomonas hydrolytica sp. DSWY01T. The molecular masses of PCLase I and PCLase II were determined to be 27.5 and 30.0 kDa, respectively. The optimum temperatures for the enzyme activities were 50 and 40°C, and the optimum pH values were 9.0 and 10.0, respectively. The two enzymes exhibited different physical and chemical properties, but both enzymes could degrade PCL substrates into monomers and oligomers. Weight loss detection and scanning electron microscopy revealed that PCLase I had more effective degradation ability than PCLase II. The genes of the two enzymes were cloned on the basis of the peptide fingerprint analysis results. The sequence analysis and substrate specificity analysis results showed that PCLase I and PCLase II were cutinase and lipase, respectively. Interface activation experiment also confirmed this conclusion. Structural analysis and modeling were further performed to obtain possible insights on the mechanism. |
| first_indexed | 2024-12-20T23:01:11Z |
| format | Article |
| id | doaj.art-8b8101ce3b0a41ddb4ca9f5db24f0138 |
| institution | Directory Open Access Journal |
| issn | 2296-4185 |
| language | English |
| last_indexed | 2024-12-20T23:01:11Z |
| publishDate | 2022-03-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Bioengineering and Biotechnology |
| spelling | doaj.art-8b8101ce3b0a41ddb4ca9f5db24f01382022-12-21T19:24:00ZengFrontiers Media S.A.Frontiers in Bioengineering and Biotechnology2296-41852022-03-011010.3389/fbioe.2022.835847835847Two Extracellular Poly(ε-caprolactone)-Degrading Enzymes From Pseudomonas hydrolytica sp. DSWY01T: Purification, Characterization, and Gene AnalysisLinying Li0Linying Li1Xiumei Lin2Jianfeng Bao3Hongmei Xia4Hongmei Xia5Fan Li6Fan Li7School of Life Sciences, Northeast Normal University, Changchun, ChinaEngineering Research Center of Glycoconjugates, Ministry of Education, Changchun, ChinaChangchun GeneScience Pharmaceutical Co., Ltd., Changchun, ChinaSchool of Life Sciences, Northeast Normal University, Changchun, ChinaSchool of Life Sciences, Northeast Normal University, Changchun, ChinaNational Demonstration Center for Experimental Biology Education, Northeast Normal University, Changchun, ChinaSchool of Life Sciences, Northeast Normal University, Changchun, ChinaEngineering Research Center of Glycoconjugates, Ministry of Education, Changchun, ChinaPoly(ε-caprolactone) (PCL) is an artificial polyester with commercially promising application. In this study, two novel PCL-degrading enzymes named PCLase I and PCLase II were purified to homogeneity from the culture supernatant of an effective polyester-degrading bacterium, Pseudomonas hydrolytica sp. DSWY01T. The molecular masses of PCLase I and PCLase II were determined to be 27.5 and 30.0 kDa, respectively. The optimum temperatures for the enzyme activities were 50 and 40°C, and the optimum pH values were 9.0 and 10.0, respectively. The two enzymes exhibited different physical and chemical properties, but both enzymes could degrade PCL substrates into monomers and oligomers. Weight loss detection and scanning electron microscopy revealed that PCLase I had more effective degradation ability than PCLase II. The genes of the two enzymes were cloned on the basis of the peptide fingerprint analysis results. The sequence analysis and substrate specificity analysis results showed that PCLase I and PCLase II were cutinase and lipase, respectively. Interface activation experiment also confirmed this conclusion. Structural analysis and modeling were further performed to obtain possible insights on the mechanism.https://www.frontiersin.org/articles/10.3389/fbioe.2022.835847/fullpoly(ε-caprolactone)Pseudomonas hydrolyticaPCL-degrading enzymecutinaselipase |
| spellingShingle | Linying Li Linying Li Xiumei Lin Jianfeng Bao Hongmei Xia Hongmei Xia Fan Li Fan Li Two Extracellular Poly(ε-caprolactone)-Degrading Enzymes From Pseudomonas hydrolytica sp. DSWY01T: Purification, Characterization, and Gene Analysis Frontiers in Bioengineering and Biotechnology poly(ε-caprolactone) Pseudomonas hydrolytica PCL-degrading enzyme cutinase lipase |
| title | Two Extracellular Poly(ε-caprolactone)-Degrading Enzymes From Pseudomonas hydrolytica sp. DSWY01T: Purification, Characterization, and Gene Analysis |
| title_full | Two Extracellular Poly(ε-caprolactone)-Degrading Enzymes From Pseudomonas hydrolytica sp. DSWY01T: Purification, Characterization, and Gene Analysis |
| title_fullStr | Two Extracellular Poly(ε-caprolactone)-Degrading Enzymes From Pseudomonas hydrolytica sp. DSWY01T: Purification, Characterization, and Gene Analysis |
| title_full_unstemmed | Two Extracellular Poly(ε-caprolactone)-Degrading Enzymes From Pseudomonas hydrolytica sp. DSWY01T: Purification, Characterization, and Gene Analysis |
| title_short | Two Extracellular Poly(ε-caprolactone)-Degrading Enzymes From Pseudomonas hydrolytica sp. DSWY01T: Purification, Characterization, and Gene Analysis |
| title_sort | two extracellular poly ε caprolactone degrading enzymes from pseudomonas hydrolytica sp dswy01t purification characterization and gene analysis |
| topic | poly(ε-caprolactone) Pseudomonas hydrolytica PCL-degrading enzyme cutinase lipase |
| url | https://www.frontiersin.org/articles/10.3389/fbioe.2022.835847/full |
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