Effect of Protein Denaturation and Enzyme Inhibitors on Proteasomal-Mediated Production of Peptides in Human Embryonic Kidney Cells

Sayani Dasgupta, Michael A. Fishman, Leandro M. Castro, Alexandre K. Tashima, Emer S. Ferro, Lloyd D. Fricker

Research output: Contribution to journalArticle

Abstract

Peptides produced by the proteasome have been proposed to function as signaling molecules that regulate a number of biological processes. In the current study, we used quantitative peptidomics to test whether conditions that affect protein stability, synthesis, or turnover cause changes in the levels of peptides in Human Embryonic Kidney 293T (HEK293T) cells. Mild heat shock (42 °C for 1 h) or treatment with the deubiquitinase inhibitor b-AP15 led to higher levels of ubiquitinated proteins but did not significantly increase the levels of intracellular peptides. Treatment with cycloheximide, an inhibitor of protein translation, did not substantially alter the levels of intracellular peptides identified herein. Cells treated with a combination of epoxomicin and bortezomib showed large increases in the levels of most peptides, relative to the levels in cells treated with either compound alone. Taken together with previous studies, these results support a mechanism in which the proteasome cleaves proteins into peptides that are readily detected in our assays (i.e., 6-37 amino acids) and then further degrades many of these peptides into smaller fragments.

Original languageEnglish (US)
JournalBiomolecules
Volume9
Issue number6
DOIs
StatePublished - May 28 2019

Fingerprint

Protein Denaturation
Denaturation
Enzyme Inhibitors
Kidney
Peptides
Proteins
Proteasome Endopeptidase Complex
Ubiquitinated Proteins
Biological Phenomena
Protein Stability
HEK293 Cells
Protein Biosynthesis
Cycloheximide
Shock
Assays
Hot Temperature
Amino Acids
Molecules

Keywords

  • bortezomib
  • epoxomicin
  • mass spectrometry
  • peptide
  • peptidomics
  • Proteasome

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology

Cite this

Effect of Protein Denaturation and Enzyme Inhibitors on Proteasomal-Mediated Production of Peptides in Human Embryonic Kidney Cells. / Dasgupta, Sayani; Fishman, Michael A.; Castro, Leandro M.; Tashima, Alexandre K.; Ferro, Emer S.; Fricker, Lloyd D.

In: Biomolecules, Vol. 9, No. 6, 28.05.2019.

Research output: Contribution to journalArticle

Dasgupta, Sayani ; Fishman, Michael A. ; Castro, Leandro M. ; Tashima, Alexandre K. ; Ferro, Emer S. ; Fricker, Lloyd D. / Effect of Protein Denaturation and Enzyme Inhibitors on Proteasomal-Mediated Production of Peptides in Human Embryonic Kidney Cells. In: Biomolecules. 2019 ; Vol. 9, No. 6.
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