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Production and statistical optimization of cholesterol-oxidase generated by Streptomyces sp. AN strain

BACKGROUND: Cholesterol oxidases (CHOs) have attracted enormous attention because of their wide biotechnological potential. The present study explores the production of CHOs by Streptomyces sp. AN. Evaluation of culture conditions affecting enzyme production, medium optimization and released metabol...

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Autores principales: Alam, Amany A., Goda, Doaa A., Soliman, Nadia A., Abdel-Meguid, Dina I., El-Sharouny, Ebaa E., Sabry, Soraya A.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Springer Berlin Heidelberg 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9649583/
https://www.ncbi.nlm.nih.gov/pubmed/36355232
http://dx.doi.org/10.1186/s43141-022-00433-1
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author Alam, Amany A.
Goda, Doaa A.
Soliman, Nadia A.
Abdel-Meguid, Dina I.
El-Sharouny, Ebaa E.
Sabry, Soraya A.
author_facet Alam, Amany A.
Goda, Doaa A.
Soliman, Nadia A.
Abdel-Meguid, Dina I.
El-Sharouny, Ebaa E.
Sabry, Soraya A.
author_sort Alam, Amany A.
collection PubMed
description BACKGROUND: Cholesterol oxidases (CHOs) have attracted enormous attention because of their wide biotechnological potential. The present study explores the production of CHOs by Streptomyces sp. AN. Evaluation of culture conditions affecting enzyme production, medium optimization and released metabolite characteristics were also investigated. RESULTS: The current work reports the isolation of 37 colonies (bacteria/actinobacteria) with different morphotypes from different soil/water samples. The isolate-coded AN was selected for its high potency for CHO production. Morphological characteristics and the obtained partial sequence of 16srRNA of AN showed 99.38% identity to Streptomyces sp. strain P12–37. Factors affecting CHO production were evaluated using Plackett-Burman (PB) and Box-Behnken (BB) statistical designs to find out the optimum level of the most effective variables, namely, pH, starch, NH(4)NO(3) and FeSO(4).7H(2)O with a predicted activity of 6.56 U/mL. According to this optimization, the following medium composition was considered to be optimum (g/L): cholesterol 1, starch 6, MgSO(4).7H(2)O 0.1, CaCl(2) 0.01, FeSO(4).7H(2)O 0.1, NH(4)NO(3) 23.97, yeast extract (YE) 0.2, K(2)HPO(4) 0.01, KH(2)PO(4) 0.1, NaCl 0.01, Tween 20 0.01, pH 6.36 and incubation temperature (30 °C) for 9 days. Spectophotometric analysis for released metabolites against cholesterol (standard) via Fourier-transform infrared spectroscopy (FTIR) and differential scanning calorimetry (DSC) was carried out. FTIR spectrum showed the appearance of new absorption peaks at 1644 and 1725cm(−1); this confirmed the presence of the Keto group (C=O) stretch bond. Besides, fermentation caused changes in thermal properties such as melting temperature peak (99.26; 148.77 °C), heat flow (− 8; − 3.6 Mw/mg), capacity (− 924.69; − 209.77 mJ) and heat enthalpy (− 385.29; 69.83 J/g) by comparison to the standard cholesterol as recognized through DSC thermogram. These changes are attributed to the action of the CHO enzyme and the release of keto derivatives of cholesterol with different properties. CONCLUSION: Streptomyces sp. AN was endowed with the capability to produce CHO. Enzyme maximization was followed using a statistical experimental approach, leading to a 2.6-fold increase in the overall activity compared to the basal condition. CHO catalyzed the oxidation of cholesterol; this was verified by the appearance of a new keto group (C=O) peak at 1644 and 1725 cm(−1) observed by FTIR spectroscopic analysis. Also, DSC thermogram demonstrates the alteration of cholesterol triggered by CHO.
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spelling pubmed-96495832022-11-28 Production and statistical optimization of cholesterol-oxidase generated by Streptomyces sp. AN strain Alam, Amany A. Goda, Doaa A. Soliman, Nadia A. Abdel-Meguid, Dina I. El-Sharouny, Ebaa E. Sabry, Soraya A. J Genet Eng Biotechnol Research BACKGROUND: Cholesterol oxidases (CHOs) have attracted enormous attention because of their wide biotechnological potential. The present study explores the production of CHOs by Streptomyces sp. AN. Evaluation of culture conditions affecting enzyme production, medium optimization and released metabolite characteristics were also investigated. RESULTS: The current work reports the isolation of 37 colonies (bacteria/actinobacteria) with different morphotypes from different soil/water samples. The isolate-coded AN was selected for its high potency for CHO production. Morphological characteristics and the obtained partial sequence of 16srRNA of AN showed 99.38% identity to Streptomyces sp. strain P12–37. Factors affecting CHO production were evaluated using Plackett-Burman (PB) and Box-Behnken (BB) statistical designs to find out the optimum level of the most effective variables, namely, pH, starch, NH(4)NO(3) and FeSO(4).7H(2)O with a predicted activity of 6.56 U/mL. According to this optimization, the following medium composition was considered to be optimum (g/L): cholesterol 1, starch 6, MgSO(4).7H(2)O 0.1, CaCl(2) 0.01, FeSO(4).7H(2)O 0.1, NH(4)NO(3) 23.97, yeast extract (YE) 0.2, K(2)HPO(4) 0.01, KH(2)PO(4) 0.1, NaCl 0.01, Tween 20 0.01, pH 6.36 and incubation temperature (30 °C) for 9 days. Spectophotometric analysis for released metabolites against cholesterol (standard) via Fourier-transform infrared spectroscopy (FTIR) and differential scanning calorimetry (DSC) was carried out. FTIR spectrum showed the appearance of new absorption peaks at 1644 and 1725cm(−1); this confirmed the presence of the Keto group (C=O) stretch bond. Besides, fermentation caused changes in thermal properties such as melting temperature peak (99.26; 148.77 °C), heat flow (− 8; − 3.6 Mw/mg), capacity (− 924.69; − 209.77 mJ) and heat enthalpy (− 385.29; 69.83 J/g) by comparison to the standard cholesterol as recognized through DSC thermogram. These changes are attributed to the action of the CHO enzyme and the release of keto derivatives of cholesterol with different properties. CONCLUSION: Streptomyces sp. AN was endowed with the capability to produce CHO. Enzyme maximization was followed using a statistical experimental approach, leading to a 2.6-fold increase in the overall activity compared to the basal condition. CHO catalyzed the oxidation of cholesterol; this was verified by the appearance of a new keto group (C=O) peak at 1644 and 1725 cm(−1) observed by FTIR spectroscopic analysis. Also, DSC thermogram demonstrates the alteration of cholesterol triggered by CHO. Springer Berlin Heidelberg 2022-11-10 /pmc/articles/PMC9649583/ /pubmed/36355232 http://dx.doi.org/10.1186/s43141-022-00433-1 Text en © The Author(s) 2022 https://creativecommons.org/licenses/by/4.0/Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) .
spellingShingle Research
Alam, Amany A.
Goda, Doaa A.
Soliman, Nadia A.
Abdel-Meguid, Dina I.
El-Sharouny, Ebaa E.
Sabry, Soraya A.
Production and statistical optimization of cholesterol-oxidase generated by Streptomyces sp. AN strain
title Production and statistical optimization of cholesterol-oxidase generated by Streptomyces sp. AN strain
title_full Production and statistical optimization of cholesterol-oxidase generated by Streptomyces sp. AN strain
title_fullStr Production and statistical optimization of cholesterol-oxidase generated by Streptomyces sp. AN strain
title_full_unstemmed Production and statistical optimization of cholesterol-oxidase generated by Streptomyces sp. AN strain
title_short Production and statistical optimization of cholesterol-oxidase generated by Streptomyces sp. AN strain
title_sort production and statistical optimization of cholesterol-oxidase generated by streptomyces sp. an strain
topic Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9649583/
https://www.ncbi.nlm.nih.gov/pubmed/36355232
http://dx.doi.org/10.1186/s43141-022-00433-1
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