Fig. 5: Characterization of RBBP9 aminopeptidase activity.
From: Mechanism-based traps enable protease and hydrolase substrate discovery
a, RBBP9 is specific for aromatic amino acids. The graph shows the catalytic efficiency of RBBP9 on 19 AA–AMCs relative to its catalytic efficiency on Phe–AMC. The bar graph represents the mean of n = 2 independent measurements. b, c, RBBP9 cleaves aromatic amino acids from the N terminus of peptide hormones (nociceptin or MENK). The full-length (FL) peptides and the products (DePhe1 or DeTyr1) after incubating with wild-type RBBP9 or RBBP9(S75A) were determined by mass spectrometry. Black solid line shows detection of product after incubating with WT RBBP9; brown solid line shows detection of product after incubating with RBBP9(S75A); black dashed line shows detection of full-length peptide after incubating with wild-type RBBP9; brown dashed line shows detection of full-length peptide after incubating with RBBP9(S75A). d, e, The crystal structure of RBBP9 in complex with Phe. d, surface view of RBBP9 and sphere representation of Phe (purple). The surface of Tyr99, Leu103, Phe140 and Leu141 is shown in yellow. e, Ribbon diagram of RBBP9 with key residues shown as sticks. Side chains of Tyr99, Leu103, Phe140 and Leu141 are shown in yellow; side chains of Glu108 and Ser76 are shown in green. The hydrogen bond between the α-amine group of Phe and the side chain of Glu108 (2.7 Å) is represented by a red dashed line. The side chain of Ser76 or Tyr99 may also form hydrogen bonds with the α-amino group of Phe (dashed brown lines, bond distances of 2.8 Å or 2.9 Å, respectively).
