Characterization of rhodanese synthesized by the wild and EMS-mutated Klebsiella oxytoca JCM1665
Cyanide poisoning remains a significant concern for both civilian and military personnel worldwide. Rhodanese, a cyanide detoxify enzyme produced by the wild (KOJCM1665) and two selected mutant strains (KOJCM1665c and KOJCM1665d) of Klebsiella oxytoca JCM 1665 was purified and characterized using a...
Saved in:
| Main Authors: | , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Elsevier
2024-12-01
|
| Series: | Journal of Agriculture and Food Research |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2666154324003958 |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| _version_ | 1846121241023348736 |
|---|---|
| author | Babamotemi Oluwasola Itakorode Oladayo Emmanuel Apalowo Isaac Duah Boateng Raphael Emuebie Okonji |
| author_facet | Babamotemi Oluwasola Itakorode Oladayo Emmanuel Apalowo Isaac Duah Boateng Raphael Emuebie Okonji |
| author_sort | Babamotemi Oluwasola Itakorode |
| collection | DOAJ |
| description | Cyanide poisoning remains a significant concern for both civilian and military personnel worldwide. Rhodanese, a cyanide detoxify enzyme produced by the wild (KOJCM1665) and two selected mutant strains (KOJCM1665c and KOJCM1665d) of Klebsiella oxytoca JCM 1665 was purified and characterized using a combination of standard techniques. Extracellular rhodanese yield of the wild and selected mutants were 6.2 ± 0.6, 26.7 ± 0.7 and 18.3 ± 0.3 U/mL respectively. The overall levels of recovery of rhodanese activity and fold after purification were 49 %, 66 %, and 58 % and 2.6, 2.6, and 2.3 % for the wild, KOJCM 1665c, and KOJCM 1665d, respectively. The native molecular weights of the three enzyme preparations were 35.1, 34.9, and 34.8 kDa, while the subunit molecular weight was 35 kDa for all the enzymes. The optimum activity of the enzymes was observed at 50 °C and pH 6.0. The real km of the three enzyme preparations for KCN as a substrate were 1.03 × 10−3, 0.95 × 10−3, and 0.80 × 10−3 M, respectively, while the km for the second substrate (Na2S2O3) were 0.59 × 10−3, 0.49 × 10−3, and 0.69 × 10−3 M, respectively. The substrate specificity study showed that the enzyme preferred sodium thiosulfate as the substrate. Metal ions such as Na+ and K+ had significantly greater inhibitory effects on enzyme activity. This study demonstrated the potential of enhancing K. oxytoca JCM 1665 to overexpress extracellular rhodanese, and the physicochemical properties of the enzyme hold significant promise as a target for improving the efficiency of cyanide bioremediation processes. |
| format | Article |
| id | doaj-art-5c590a3bf5cf45fd9b9f79d6224a204c |
| institution | Kabale University |
| issn | 2666-1543 |
| language | English |
| publishDate | 2024-12-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Journal of Agriculture and Food Research |
| spelling | doaj-art-5c590a3bf5cf45fd9b9f79d6224a204c2024-12-16T05:37:31ZengElsevierJournal of Agriculture and Food Research2666-15432024-12-0118101358Characterization of rhodanese synthesized by the wild and EMS-mutated Klebsiella oxytoca JCM1665Babamotemi Oluwasola Itakorode0Oladayo Emmanuel Apalowo1Isaac Duah Boateng2Raphael Emuebie Okonji3Department of Biotechnology, Osun State University, Osogbo, Osun State, Nigeria; Corresponding author.Department of Biochemistry, Nutrition and Health Promotion, Mississippi State University, Starkville, MS, USAOrganization of African Academic Doctors, PO Box 25305-00100, Nairobi, Kenya; Certified Group, 199 W Rhapsody Dr, San Antonio, TX, TX, 78216, USADepartment of Biochemistry and Molecular Biology, Obafemi Awolowo University, Ile-Ife, Osun State, NigeriaCyanide poisoning remains a significant concern for both civilian and military personnel worldwide. Rhodanese, a cyanide detoxify enzyme produced by the wild (KOJCM1665) and two selected mutant strains (KOJCM1665c and KOJCM1665d) of Klebsiella oxytoca JCM 1665 was purified and characterized using a combination of standard techniques. Extracellular rhodanese yield of the wild and selected mutants were 6.2 ± 0.6, 26.7 ± 0.7 and 18.3 ± 0.3 U/mL respectively. The overall levels of recovery of rhodanese activity and fold after purification were 49 %, 66 %, and 58 % and 2.6, 2.6, and 2.3 % for the wild, KOJCM 1665c, and KOJCM 1665d, respectively. The native molecular weights of the three enzyme preparations were 35.1, 34.9, and 34.8 kDa, while the subunit molecular weight was 35 kDa for all the enzymes. The optimum activity of the enzymes was observed at 50 °C and pH 6.0. The real km of the three enzyme preparations for KCN as a substrate were 1.03 × 10−3, 0.95 × 10−3, and 0.80 × 10−3 M, respectively, while the km for the second substrate (Na2S2O3) were 0.59 × 10−3, 0.49 × 10−3, and 0.69 × 10−3 M, respectively. The substrate specificity study showed that the enzyme preferred sodium thiosulfate as the substrate. Metal ions such as Na+ and K+ had significantly greater inhibitory effects on enzyme activity. This study demonstrated the potential of enhancing K. oxytoca JCM 1665 to overexpress extracellular rhodanese, and the physicochemical properties of the enzyme hold significant promise as a target for improving the efficiency of cyanide bioremediation processes.http://www.sciencedirect.com/science/article/pii/S2666154324003958CyanideBioremediationRhodaneseBioprocessMutagenesisEnzyme kinetics |
| spellingShingle | Babamotemi Oluwasola Itakorode Oladayo Emmanuel Apalowo Isaac Duah Boateng Raphael Emuebie Okonji Characterization of rhodanese synthesized by the wild and EMS-mutated Klebsiella oxytoca JCM1665 Journal of Agriculture and Food Research Cyanide Bioremediation Rhodanese Bioprocess Mutagenesis Enzyme kinetics |
| title | Characterization of rhodanese synthesized by the wild and EMS-mutated Klebsiella oxytoca JCM1665 |
| title_full | Characterization of rhodanese synthesized by the wild and EMS-mutated Klebsiella oxytoca JCM1665 |
| title_fullStr | Characterization of rhodanese synthesized by the wild and EMS-mutated Klebsiella oxytoca JCM1665 |
| title_full_unstemmed | Characterization of rhodanese synthesized by the wild and EMS-mutated Klebsiella oxytoca JCM1665 |
| title_short | Characterization of rhodanese synthesized by the wild and EMS-mutated Klebsiella oxytoca JCM1665 |
| title_sort | characterization of rhodanese synthesized by the wild and ems mutated klebsiella oxytoca jcm1665 |
| topic | Cyanide Bioremediation Rhodanese Bioprocess Mutagenesis Enzyme kinetics |
| url | http://www.sciencedirect.com/science/article/pii/S2666154324003958 |
| work_keys_str_mv | AT babamotemioluwasolaitakorode characterizationofrhodanesesynthesizedbythewildandemsmutatedklebsiellaoxytocajcm1665 AT oladayoemmanuelapalowo characterizationofrhodanesesynthesizedbythewildandemsmutatedklebsiellaoxytocajcm1665 AT isaacduahboateng characterizationofrhodanesesynthesizedbythewildandemsmutatedklebsiellaoxytocajcm1665 AT raphaelemuebieokonji characterizationofrhodanesesynthesizedbythewildandemsmutatedklebsiellaoxytocajcm1665 |