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Identification of newly translated thermo-sensitive proteins using pulse SILAC mass spectrometry and the GAL promoter system.

ABSTRACT: Some newly translated proteins are more susceptible to misfolding and aggregation upon heat shock in comparison to other proteins. To study these newly translated thermo-sensitive proteins on a proteomic scale, we present here a protocol that combines pulse-SILAC with biochemical fractionation for mass spectrometry analysis, followed by an orthogonal validation protocol for selected candidates using the GAL promoter system in Saccharomyces cerevisiae. This approach can be further developed to study other stresses and specific post-translational modifications or adapted to mammalian cells. For complete details on the use and execution of this protocol, please refer to Zhu et al. (2022).¹.

SUBMITTER: Zhu M

PROVIDER: S-EPMC9881406 | biostudies-literature | 2023 Jan

REPOSITORIES: biostudies-literature

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Identification of newly translated thermo-sensitive proteins using pulse SILAC mass spectrometry and the GAL promoter system.

Zhu Mang M Calabrese Gaetano G Wong Ryan W K RWK Mayor Thibault T

STAR protocols 20230124 1

Some newly translated proteins are more susceptible to misfolding and aggregation upon heat shock in comparison to other proteins. To study these newly translated thermo-sensitive proteins on a proteomic scale, we present here a protocol that combines pulse-SILAC with biochemical fractionation for mass spectrometry analysis, followed by an orthogonal validation protocol for selected candidates using the GAL promoter system in Saccharomyces cerevisiae. This approach can be further developed to st ...[more]

PMID: 36853680

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