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https://hdl.handle.net/11147/9600
Title: | Effects of N-terminal and C-terminal polyhistidine tag on the stability and function of the thermophilic P450 CYP119 | Authors: | Aslantaş, Yaprak Sürmeli, Nur Başak |
Issue Date: | 2019 | Publisher: | Hindawi Publishing Corporation | Abstract: | Biocatalysts are sought-after in synthesis of pharmaceuticals and agrochemicals due to their high regioselectivity and enantioselectivity. Among biocatalysts, heme-containing cytochrome P450 (P450) oxygenases are an attractive target since they catalyze oxidation of "unactivated" carbon-hydrogen bonds with high efficiency. CYP119 is an acidothermophilic P450 from Sulfolobus acidocaldarius, which has the potential to be widely used as a biocatalyst since it shows activity at high temperatures and low pH. Polyhistidine tags (His-tags) are widely used to simplify purification of proteins. However, His-tags can cause changes to protein structure and function. Here, we demonstrate the effects of His-tags on CYP119. To this end, the His-tags were cloned at the N-terminus or C-terminus of the CYP119, and His-tagged proteins were expressed and isolated. The thermostability and peroxidase activity of His-tagged CYP119s were tested and compared to wild type CYP119. Results indicated that while addition of His-tags increased the yield and simplified isolation of CYP119, they also influenced the electronic structure of active site and the activity of the protein. We show that N-terminal His-tagged CYP119 has desirable properties and potential to be used in industrial applications, but mechanistic studies using this protein need careful interpretation since the His-tag affects electronic properties of the active site heme iron. | URI: | https://doi.org/10.1155/2019/8080697 https://hdl.handle.net/11147/9600 |
ISSN: | 1565-3633 1687-479X |
Appears in Collections: | Bioengineering / Biyomühendislik PubMed İndeksli Yayınlar Koleksiyonu / PubMed Indexed Publications Collection Scopus İndeksli Yayınlar Koleksiyonu / Scopus Indexed Publications Collection WoS İndeksli Yayınlar Koleksiyonu / WoS Indexed Publications Collection |
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8080697.pdf | 1.79 MB | Adobe PDF | View/Open |
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