Comprehensive analysis of LIR binding interactions with Atg8-family proteins, FIP200 ^Claw , and TAX1BP1 ^CC2 . (A) The LIR peptide array, containing peptides selected from the iLIR database, was probed with 100 nM GST-LC3A, GST-GABARAPL1, GST-FIP200^Claw, or TAX1BP1^CC2-MBP. Antibody-HRP conjugates were used for visualization. Peptide spots that bound none of the four protein probes (“never-binders”) are labeled with red asterisk and excluded from further analysis. Redundant protein names reflect multiple LIR sequences within a single protein. LIR motif of each peptide is shown for reference. (B) FIR containing peptides from selected literature, probed as in A. (C) Spearman r correlation analysis of SLiM binding preference for each protein (LC3A, GABARAPL1, GST-FIP200^Claw, or TAX1BP1^CC2-MBP) based on spot intensity in A and B. “Never-binders” and phosphomimetic peptides were excluded from analysis. N = 79. (D) IceLogo sequence alignment derived from all bound peptides (determined by blinded scorers). All unbound peptides were used as a reference set to calculate position specific P value for each amino acid. “Never-binders” were excluded from analysis. Enriched amino acids are positive, under-represented amino acids are negative. Shown as percent enrichment (P value <0.05).