Evidence of excited-state lifetime enhancement in dimyristoyl-phosphocholine nanodiscs by ultraviolet absorption spectroscopy
Ultraviolet absorption spectroscopy is an analytical tool that is commonly utilized to determine protein concentrations, primarily due to characteristic absorption of tryptophan and tyrosine at 280 nm. Accurate concentration determination is essential to the accuracy of many biophysical techniques,...
| Main Authors: | , , |
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| Format: | Article |
| Language: | English |
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AIP Publishing LLC
2023-01-01
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| Series: | AIP Advances |
| Online Access: | http://dx.doi.org/10.1063/5.0102149 |
| _version_ | 1811172594251988992 |
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| author | Philip A. Kocheril Miranda I. Hiller Jessica Z. Kubicek-Sutherland |
| author_facet | Philip A. Kocheril Miranda I. Hiller Jessica Z. Kubicek-Sutherland |
| author_sort | Philip A. Kocheril |
| collection | DOAJ |
| description | Ultraviolet absorption spectroscopy is an analytical tool that is commonly utilized to determine protein concentrations, primarily due to characteristic absorption of tryptophan and tyrosine at 280 nm. Accurate concentration determination is essential to the accuracy of many biophysical techniques, and ultraviolet absorption provides a convenient and rapid method to assess protein concentration. However, the widespread usage of this method assumes that ultraviolet absorption is the same for individual amino acids in aqueous solution (where the molar extinction coefficient was measured) as for the amino acid in the conformational environment of the protein of interest. We demonstrate that additional considerations may be necessary for the membrane scaffold protein in dimyristoyl-phosphocholine nanodiscs through ultraviolet absorption spectroscopy of nanodiscs, liposomes, the membrane scaffold protein, and a mixture of liposomes and the non-nanodisc-associated protein. The aromatic amino acids of the membrane scaffold protein absorb significantly less light at 280 nm when associated with lipids in a lipoprotein assembly, which we plausibly attribute to an enhancement of chromophore excited-state lifetimes due to reduced intramolecular motion in the nanodisc. We caution that using the molar absorptivity of the membrane scaffold protein alone to determine nanodisc concentrations may not yield accurate results. Instead, the molar absorptivity of each nanodisc formulation should be explored independently to account for the unique conformational environment of each nanodisc. |
| first_indexed | 2024-04-10T17:33:53Z |
| format | Article |
| id | doaj.art-c7735b91547b4729844722f34429f17f |
| institution | Directory Open Access Journal |
| issn | 2158-3226 |
| language | English |
| last_indexed | 2024-04-10T17:33:53Z |
| publishDate | 2023-01-01 |
| publisher | AIP Publishing LLC |
| record_format | Article |
| series | AIP Advances |
| spelling | doaj.art-c7735b91547b4729844722f34429f17f2023-02-03T16:42:07ZengAIP Publishing LLCAIP Advances2158-32262023-01-01131015124015124-610.1063/5.0102149Evidence of excited-state lifetime enhancement in dimyristoyl-phosphocholine nanodiscs by ultraviolet absorption spectroscopyPhilip A. Kocheril0Miranda I. Hiller1Jessica Z. Kubicek-Sutherland2Physical Chemistry and Applied Spectroscopy, Chemistry Division, Los Alamos National Laboratory, Los Alamos, New Mexico 87545, USAPhysical Chemistry and Applied Spectroscopy, Chemistry Division, Los Alamos National Laboratory, Los Alamos, New Mexico 87545, USAPhysical Chemistry and Applied Spectroscopy, Chemistry Division, Los Alamos National Laboratory, Los Alamos, New Mexico 87545, USAUltraviolet absorption spectroscopy is an analytical tool that is commonly utilized to determine protein concentrations, primarily due to characteristic absorption of tryptophan and tyrosine at 280 nm. Accurate concentration determination is essential to the accuracy of many biophysical techniques, and ultraviolet absorption provides a convenient and rapid method to assess protein concentration. However, the widespread usage of this method assumes that ultraviolet absorption is the same for individual amino acids in aqueous solution (where the molar extinction coefficient was measured) as for the amino acid in the conformational environment of the protein of interest. We demonstrate that additional considerations may be necessary for the membrane scaffold protein in dimyristoyl-phosphocholine nanodiscs through ultraviolet absorption spectroscopy of nanodiscs, liposomes, the membrane scaffold protein, and a mixture of liposomes and the non-nanodisc-associated protein. The aromatic amino acids of the membrane scaffold protein absorb significantly less light at 280 nm when associated with lipids in a lipoprotein assembly, which we plausibly attribute to an enhancement of chromophore excited-state lifetimes due to reduced intramolecular motion in the nanodisc. We caution that using the molar absorptivity of the membrane scaffold protein alone to determine nanodisc concentrations may not yield accurate results. Instead, the molar absorptivity of each nanodisc formulation should be explored independently to account for the unique conformational environment of each nanodisc.http://dx.doi.org/10.1063/5.0102149 |
| spellingShingle | Philip A. Kocheril Miranda I. Hiller Jessica Z. Kubicek-Sutherland Evidence of excited-state lifetime enhancement in dimyristoyl-phosphocholine nanodiscs by ultraviolet absorption spectroscopy AIP Advances |
| title | Evidence of excited-state lifetime enhancement in dimyristoyl-phosphocholine nanodiscs by ultraviolet absorption spectroscopy |
| title_full | Evidence of excited-state lifetime enhancement in dimyristoyl-phosphocholine nanodiscs by ultraviolet absorption spectroscopy |
| title_fullStr | Evidence of excited-state lifetime enhancement in dimyristoyl-phosphocholine nanodiscs by ultraviolet absorption spectroscopy |
| title_full_unstemmed | Evidence of excited-state lifetime enhancement in dimyristoyl-phosphocholine nanodiscs by ultraviolet absorption spectroscopy |
| title_short | Evidence of excited-state lifetime enhancement in dimyristoyl-phosphocholine nanodiscs by ultraviolet absorption spectroscopy |
| title_sort | evidence of excited state lifetime enhancement in dimyristoyl phosphocholine nanodiscs by ultraviolet absorption spectroscopy |
| url | http://dx.doi.org/10.1063/5.0102149 |
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