Cucurbit[7]uril Enhances Distance Measurements of Spin-Labeled Proteins
We report complex formation between the chloroacetamide 2,6-diazaadamantane nitroxide radical (ClA-DZD) and cucurbit[7]uril (CB-7), for which the association constant in water, Ka = 1.9 × 106 M-1, is at least 1 order of magnitude higher than the previously studied organic radicals. The radical is highly immobilized by CB-7, as indicated by the increase in the rotational correlation time, τrot, by a factor of 36, relative to that in the buffer solution. The X-ray structure of ClA-DZDCB-7 shows the encapsulated DZD guest inside the undistorted CB-7 host, with the pendant group protruding outside. Upon addition of CB-7 to T4 Lysozyme (T4L) doubly spin-labeled with the iodoacetamide derivative of DZD, we observe the increase in τrot and electron spin coherence time, Tm, along with the narrowing of interspin distance distributions. Sensitivity of the DEER measurements at 83 K increases by a factor 4-9, compared to the common spin label such as MTSL, which is not affected by CB-7. Interspin distances of 3 nm could be reliably measured in water/glycerol up to temperatures near the glass transition/melting temperature of the matrix at 200 K, thus bringing us closer to the goal of supramolecular recognition-enabled long-distance DEER measurements at near physiological temperatures. The X-ray structure of DZD-T4L 65 at 1.12 Å resolution allows for unambiguous modeling of the DZD label (0.88 occupancy), indicating an undisturbed structure and conformation of the protein.
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| Medienart: |
E-Artikel |
| Erscheinungsjahr: |
2023 |
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| Erschienen: |
2023 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:145 |
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| Enthalten in: |
Journal of the American Chemical Society - 145(2023), 47 vom: 29. Nov., Seite 25726-25736 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Yang, Zhimin [VerfasserIn] |
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| Links: |
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| Themen: |
059QF0KO0R |
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| Anmerkungen: |
Date Completed 30.11.2023 Date Revised 26.03.2024 published: Print-Electronic UpdateOf: bioRxiv. 2023 Aug 23;:. - PMID 37662277 Citation Status MEDLINE |
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| doi: |
10.1021/jacs.3c09184 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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| PPN (Katalog-ID): |
NLM36454807X |
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| 245 | 1 | 0 | |a Cucurbit[7]uril Enhances Distance Measurements of Spin-Labeled Proteins |
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| 500 | |a Date Revised 26.03.2024 | ||
| 500 | |a published: Print-Electronic | ||
| 500 | |a UpdateOf: bioRxiv. 2023 Aug 23;:. - PMID 37662277 | ||
| 500 | |a Citation Status MEDLINE | ||
| 520 | |a We report complex formation between the chloroacetamide 2,6-diazaadamantane nitroxide radical (ClA-DZD) and cucurbit[7]uril (CB-7), for which the association constant in water, Ka = 1.9 × 106 M-1, is at least 1 order of magnitude higher than the previously studied organic radicals. The radical is highly immobilized by CB-7, as indicated by the increase in the rotational correlation time, τrot, by a factor of 36, relative to that in the buffer solution. The X-ray structure of ClA-DZDCB-7 shows the encapsulated DZD guest inside the undistorted CB-7 host, with the pendant group protruding outside. Upon addition of CB-7 to T4 Lysozyme (T4L) doubly spin-labeled with the iodoacetamide derivative of DZD, we observe the increase in τrot and electron spin coherence time, Tm, along with the narrowing of interspin distance distributions. Sensitivity of the DEER measurements at 83 K increases by a factor 4-9, compared to the common spin label such as MTSL, which is not affected by CB-7. Interspin distances of 3 nm could be reliably measured in water/glycerol up to temperatures near the glass transition/melting temperature of the matrix at 200 K, thus bringing us closer to the goal of supramolecular recognition-enabled long-distance DEER measurements at near physiological temperatures. The X-ray structure of DZD-T4L 65 at 1.12 Å resolution allows for unambiguous modeling of the DZD label (0.88 occupancy), indicating an undisturbed structure and conformation of the protein | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a Research Support, N.I.H., Extramural | |
| 650 | 4 | |a Research Support, Non-U.S. Gov't | |
| 650 | 4 | |a Research Support, U.S. Gov't, Non-P.H.S. | |
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| 700 | 1 | |a Stein, Richard A |e verfasserin |4 aut | |
| 700 | 1 | |a Pink, Maren |e verfasserin |4 aut | |
| 700 | 1 | |a Madzelan, Peter |e verfasserin |4 aut | |
| 700 | 1 | |a Ngendahimana, Thacien |e verfasserin |4 aut | |
| 700 | 1 | |a Rajca, Suchada |e verfasserin |4 aut | |
| 700 | 1 | |a Wilson, Mark A |e verfasserin |4 aut | |
| 700 | 1 | |a Eaton, Sandra S |e verfasserin |4 aut | |
| 700 | 1 | |a Eaton, Gareth R |e verfasserin |4 aut | |
| 700 | 1 | |a Mchaourab, Hassane S |e verfasserin |4 aut | |
| 700 | 1 | |a Rajca, Andrzej |e verfasserin |4 aut | |
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