Figure 4. Multiple substrate residues contribute synergistically to grip.

(A) GA and Ala-4 cassette sequences. A heatmap of V_max values from Figure 2C is overlaid to show contribution of single tyrosine residues as each tail position. (B) Fraction intracellular degradation of substrates with one alanine at tail-position 4 and a second alanine at a variable position in otherwise all-glycine cassettes. (C) Comparison of intracellular degradation for a subset of substrates, including Ala-1. (D) V_max values from Michaelis-Menten analysis of degradation of purified substrates. (E and F) Michaelis-Menten V_max values for purified substrates with one tyrosine (E) or valine (F) at tail-position four and a second tyrosine (E) or valine (F) at each tail position in otherwise all-glycine cassettes. Overlaid dashed lines indicate degradation rate for the parental Tyr-4 (E) or Val-4 (F) substrates. In all panels, values represent averages (± S.D.) of three biological replicates.

Figure 4—figure supplement 1. Degradation of Dual-Tyr substrates centered at tail position 3.

V_max values for degradation from Michaelis-Menten analysis of purified substrates with one tyrosine at tail-position three and a second tyrosine at a variable position in otherwise all-glycine cassettes.

Relative degradation for substrate tails with a single Tyr residue at position 3 or 4 indicated by dashed lines. Values represent averages (± S.D.) of three biological replicates.