2023
DOI: 10.1073/pnas.2303167120
|View full text |Cite
|
Sign up to set email alerts
|

Folding stabilities of ribosome-bound nascent polypeptides probed by mass spectrometry

Abstract: The folding of most proteins occurs during the course of their translation while their tRNA-bound C termini are embedded in the ribosome. How the close proximity of nascent proteins to the ribosome influences their folding thermodynamics remains poorly understood. Here, we have developed a mass spectrometry–based approach for determining the stabilities of nascent polypeptide chains using methionine oxidation as a folding probe. This approach enables quantitative measurement subglobal folding stabilities of ri… Show more

Help me understand this report

Search citation statements

Order By: Relevance

Paper Sections

Select...
2
1
1
1

Citation Types

0
5
0

Year Published

2024
2024
2024
2024

Publication Types

Select...
4
2

Relationship

2
4

Authors

Journals

citations
Cited by 6 publications
(5 citation statements)
references
References 59 publications
0
5
0
Order By: Relevance
“…The peptide mixture was pulse oxidized by exposure to varying concentrations of unlabeled ( 16 O) hydrogen peroxide and subsequently blocked by addition of excess levels of 18 O-labeled hydrogen peroxide. As shown previously, blocking of peptides by 18 O-labeled hydrogen peroxide prevents artifactual oxidation of methionines during the subsequent bottom-up proteomics workflow 34, 35 . After blocking, fractional oxidation during the pulse phase of the experiment can be accurately quantified by determining the relative ratio of 16 O to 18 O-labeled peptides from the resulting mass spectra (Figure 2A).…”
Section: Formylation Facilitates Reduction Of Nt-methionine By Msrs I...mentioning
confidence: 79%
See 2 more Smart Citations
“…The peptide mixture was pulse oxidized by exposure to varying concentrations of unlabeled ( 16 O) hydrogen peroxide and subsequently blocked by addition of excess levels of 18 O-labeled hydrogen peroxide. As shown previously, blocking of peptides by 18 O-labeled hydrogen peroxide prevents artifactual oxidation of methionines during the subsequent bottom-up proteomics workflow 34, 35 . After blocking, fractional oxidation during the pulse phase of the experiment can be accurately quantified by determining the relative ratio of 16 O to 18 O-labeled peptides from the resulting mass spectra (Figure 2A).…”
Section: Formylation Facilitates Reduction Of Nt-methionine By Msrs I...mentioning
confidence: 79%
“…Fragpipe-supplied PSM files were used for quantitation of PSM numbers. Quantification of 16 O/ 18 O labeled methionine ratios was carried out as previously described 34, 35 . Briefly, MS1 spectra corresponding to specific methionine-containing peptides were extracted from the data.…”
Section: Lc-ms/ms Analysismentioning
confidence: 99%
See 1 more Smart Citation
“…MS1 files were generated from raw files using MSConvert 22 and fractional oxidations were measured as previously described. 19 We note that under some conditions, a small population of the peptide was oxidized to form methionine sulfones in addition to methionine sulfoxides. In these cases, fractional oxidations were measured using the population averaged ratios of 16 O/ 18 O-oxidized relative to 18 O/ 18 O-oxidized peptides, as well as 16 O-oxidized relative to 18 O-oxidized peptides.…”
Section: Mass Spectrometrymentioning
confidence: 94%
“…Relative levels of unmodified, 16 O-and 18 O-oxidized peptides were determined by analyzing the intact mass spectra as described previously. 19 To determine the concentration of H2 18 O2, 10 µg of a synthetic peptide (MASLIKKLAVDR) was incubated with differing concentrations of H2 16 O2 (Fisher Bioreagents) or a 2x or 5x dilution of the generated H2 18 O2 solution for 10 minutes at 37 °C. Oxidation was quenched with 400 mM sodium sulfite and samples were frozen and lyophilized.…”
Section: Determination Of the Purity And Concentration Of H2 18 O2mentioning
confidence: 99%