Table 1.
Statistics for data collection, phasing, and model refinement.
Figure 1.
Overview of the crystal structure of hNtaq1.
(A) Overall structure of hNtaq1. α-helices, β-strands, and loops are colored in orange, cyan, and white, respectively. The representative amino acid residues in the active site are shown as stick model (carbon, oxygen, nitrogen, and sulfur in yellow, red, blue, and gold color, respectively). (B) Stereo view of the crystallographic contact of hNtaq1 with a symmetry-related molecule. hNtaq1 and symmetry-related molecules are represented as green, cyan, and magenta cartoon, respectively. Unit cell is shown with green line and electron density map is shown as gray cloud. (C) Sequence alignment of hNtaq1 with Ntaq proteins from Mus musculus, Caenorhabditis elegans, protein-glutaminase from Chryseobacterium proteolyticum, secreted effector protein SseI from Sallonella typhimurium, and periplasmic protease LapG from Legionella pneumophila. Residues of the catalytic triad are represented by red asterisk below the amino acid sequences. Completely conserved, identical, moderately conserved residues are highlighted with red, green, and yellow shaded boxes, respectively. The secondary structure of hNtaq1 is shown on top of the sequence alignment. α-helix, β-sheet, and connecting region are represented by red spiral, blue arrow, and black line, respectively. Structural comparison of hNtaq1 with protein-glutaminase from C. proteolyticum (D), secreted effector protein SseI from S. typhimurium (E), and periplasmic protease LapG from L. peumophila (F). The catalytic triads are shown as stick models and main chains of hNtaq1, protein-glutaminase, SseI, and LapG are represented with ribbon diagram in green, magenta, orange, and blue, respectively.
Figure 2.
Active site and electrostatic potential surface charge of hNtaq1.
(A) Substrate binding cleft of hNtaq1. Carbon in the substrate-mimicking peptide, catalytic triad, α-helices, β-strands, and loops are colored in green, yellow, orange, cyan, and white, respectively. Oxygen, nitrogen, and sulfur atoms are represented as red, blue, and gold, respectively. Two water molecules are shown as red sphere and labeled as W1 and W2. (B) Electron density map from an Fo–Fc omit map calculated without the bound substrate-mimicking peptide. Positive electron density are shown as a green mesh contoured at 2.0 σ, in a stereo view. (C) Electrostatic potential surface and substrate binding cleft region of hNtaq1. Negatively and positively charged surfaces are represented as red and blue shade, respectively. Residues interacting with the substrate-mimicking peptide molecule are labeled.
Figure 3.
Docking study and suggested binding pose of N-terminal glutamine peptide in hNtaq1.
(A) Tripeptides docking study of hNtaq1. Categorized by features of its second residue, the backbone of Ala, Gly, Ile, Leu, Met, Pro, Val tripeptides are colored in yellow, the backbone of Cys, Asn, Gln, Ser, Thr tripetides are colored in green, the His, Lys, Arg tripeptides are colored in blue, the Phe, Trp, Tyr tripeptides are colored in black, and the Asp, Glu tripeptides are colored in orange. (B) The nearest tripeptide in docking study. The substrate-mimicking peptide is shown as green and predicted docking tripeptide Gln-Tyr-Pro is colored in magenta. (C) Binding mode of refined Ser1Gln hNtaq1 mutant on electron density map of hNtaq1. Carbon in substrate-mimicking peptide, catalytic triad, β-strands, loops, and Ser1Gln mutant are colored in green, yellow, cyan, white, and blue, respectively. Oxygen, nitrogen, and sulfur are represented as red, blue, and gold, respectively. Two water molecules are shown as red sphere and electron density map is represented as gray mesh contoured at 2.0 σ.
Figure 4.
Proposed catalytic mechanism of hNtaq1.