Figure 3.

Amino acid frequency at flanking positions of di-lysine and arginine-based signals. (A) Frequency plots for each randomized position in the di-lysine signal libraries. Unique sequences identified for both libraries were analyzed with respect to the amino acid frequency at each randomized position in the pool of intracellularly retained (x axis) or surface-expressed (y axis) reporter proteins. The number of unique sequences were 56 (-KKXX-C, retained), 48 (-KKXX-C, surface), 39 (-XXXKTN-C, retained) and 39 (-XXXKTN, surface). The position analyzed in the respective plot is indicated in green. Amino acids favored in the surface pool are indicated in red, whereas amino acids favored in the retained pool are shown in blue. Amino acids shown in black were either rare or equally abundant in both pools. (B) Table of clones selected from the library of arginine-based signal variants. Blue randomized positions indicated ER localization, whereas red randomized positions indicate surface expression. Tables 1 and 2 (which are published as supplementary material on the PNAS web site, www.pnas.org) list the original sequences obtained from the screen.