Table 3 Peptides

From: Mechanism of millisecond Lys48-linked poly-ubiquitin chain formation by cullin-RING ligases

Peptide

Sequence

Source

Cyclin E ‘sortasing’ assays

GGGGPLPAGLL(pT)PPQ(pS)GRRASY

21

Cyclin E ‘sortasing’ cryo-EM

GGGGLPSGLL(pT)PPQ(pS)GKKQSSDYKDDDDK

21

Cyclin E substrate assays

Ac-KAMLSEQNRASPLPSGLL(pT)PPQ(pS)GRRASY

21

β-catenin ‘sortasing’ assays

GGGGYLD(pS)GIH(pS)GATTAPRRASY

22

Hif1α ‘sortasing’ assays

GGGGLLA(hyP)PAAGDTIISLDFGSNGRRASY

MPI

Hif1α substrate assays

Ac-KLRREPDALTLLA(hyP)AAGDTIISLDFGSN-Fluorescein

MPI

Sil1 substrate assays

Ac-GRRASYGSGSKEGYFQELLGSVNPTQGRAR

NEP

  1. All peptides were either purchased from Vivitide (formerly New England Peptides (NEP); greater than 95% purity) or synthesized in-house at the Max Planck Institute of Biochemistry (MPI) and solubilized in water. All single lysine peptide substrates had their N termini acetylated (Ac). Phosphodegrons are shown as pT (phospho Thr) or pS (phospho Ser), and the hydroxylated Pro degron in Hif1α peptides are shown as hyP. The Sil1 peptide substrate amino acid sequence was based on the clone 13 design17 that had optimized affinity for FEM1C. All peptides that were substrates for ubiquitylation assays contained the ‘RRASY’ amino acidic sequence that enabled 32P-labeling by protein kinase A (New England Biolabs).
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