Substrates incorporating unnatural amino acids are separately detectable for chymotrypsin and subtilisin with similar specificity
Protease specificity depends on the three-dimensional structure and sequence of the protease, and each protease plays a different role. Chymotrypsin and subtilisin have similar specificity for recognizing aromatic and hydrophobic amino acid residues. Previous methods using natural amino acids have d...
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| Format: | Article |
| Language: | English |
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Elsevier
2023-12-01
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| Series: | Results in Chemistry |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2211715623004563 |
| _version_ | 1797398666977214464 |
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| author | Yuuki Yamawaki Tamaki Kato |
| author_facet | Yuuki Yamawaki Tamaki Kato |
| author_sort | Yuuki Yamawaki |
| collection | DOAJ |
| description | Protease specificity depends on the three-dimensional structure and sequence of the protease, and each protease plays a different role. Chymotrypsin and subtilisin have similar specificity for recognizing aromatic and hydrophobic amino acid residues. Previous methods using natural amino acids have difficulty in separately detecting proteases with similar specificity. To overcome this, we developed an effective substrate incorporating unnatural amino acids using a simple method that changes only one residue of a substrate. Various 7-amino-4-methylcoumarin (AMC) peptide substrates were synthesized in terms of hydrophobicity, aromaticity, and conformation, including unnatural amino acids 2-amino-4-ethylhexanoic acid (AEH) and cyclohexylalanine (Cha). Although AEH substrate is highly hydrophobic, steric hindrance based on the branched side chain of AEH residue reduced the recognition of the substrate by the two proteases. Cha substrate with a six-membered ring side chain was most preferably recognized by chymotrypsin but was slightly recognized by subtilisin. This shows that Cha substrate can separately detect the two proteases with similar specificity. Therefore, these results indicate that protease with similar specificities is separately detectable by substrates with a side chain having structural degrees of freedom that provide different steric hindrance. |
| first_indexed | 2024-03-09T01:28:53Z |
| format | Article |
| id | doaj.art-f4a2f5fbb0114c03a910d7d36ddb234e |
| institution | Directory Open Access Journal |
| issn | 2211-7156 |
| language | English |
| last_indexed | 2024-03-09T01:28:53Z |
| publishDate | 2023-12-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Results in Chemistry |
| spelling | doaj.art-f4a2f5fbb0114c03a910d7d36ddb234e2023-12-10T06:15:40ZengElsevierResults in Chemistry2211-71562023-12-016101217Substrates incorporating unnatural amino acids are separately detectable for chymotrypsin and subtilisin with similar specificityYuuki Yamawaki0Tamaki Kato1Graduate School of Life Science and Systems Engineering, Kyushu Institute of Technology, 2-4 Hibikino, Wakamatsu-ku, Kitakyushu, Fukuoka 808-0196, JapanCorresponding author.; Graduate School of Life Science and Systems Engineering, Kyushu Institute of Technology, 2-4 Hibikino, Wakamatsu-ku, Kitakyushu, Fukuoka 808-0196, JapanProtease specificity depends on the three-dimensional structure and sequence of the protease, and each protease plays a different role. Chymotrypsin and subtilisin have similar specificity for recognizing aromatic and hydrophobic amino acid residues. Previous methods using natural amino acids have difficulty in separately detecting proteases with similar specificity. To overcome this, we developed an effective substrate incorporating unnatural amino acids using a simple method that changes only one residue of a substrate. Various 7-amino-4-methylcoumarin (AMC) peptide substrates were synthesized in terms of hydrophobicity, aromaticity, and conformation, including unnatural amino acids 2-amino-4-ethylhexanoic acid (AEH) and cyclohexylalanine (Cha). Although AEH substrate is highly hydrophobic, steric hindrance based on the branched side chain of AEH residue reduced the recognition of the substrate by the two proteases. Cha substrate with a six-membered ring side chain was most preferably recognized by chymotrypsin but was slightly recognized by subtilisin. This shows that Cha substrate can separately detect the two proteases with similar specificity. Therefore, these results indicate that protease with similar specificities is separately detectable by substrates with a side chain having structural degrees of freedom that provide different steric hindrance.http://www.sciencedirect.com/science/article/pii/S2211715623004563Protease activity |
| spellingShingle | Yuuki Yamawaki Tamaki Kato Substrates incorporating unnatural amino acids are separately detectable for chymotrypsin and subtilisin with similar specificity Results in Chemistry Protease activity |
| title | Substrates incorporating unnatural amino acids are separately detectable for chymotrypsin and subtilisin with similar specificity |
| title_full | Substrates incorporating unnatural amino acids are separately detectable for chymotrypsin and subtilisin with similar specificity |
| title_fullStr | Substrates incorporating unnatural amino acids are separately detectable for chymotrypsin and subtilisin with similar specificity |
| title_full_unstemmed | Substrates incorporating unnatural amino acids are separately detectable for chymotrypsin and subtilisin with similar specificity |
| title_short | Substrates incorporating unnatural amino acids are separately detectable for chymotrypsin and subtilisin with similar specificity |
| title_sort | substrates incorporating unnatural amino acids are separately detectable for chymotrypsin and subtilisin with similar specificity |
| topic | Protease activity |
| url | http://www.sciencedirect.com/science/article/pii/S2211715623004563 |
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