The three-dimensional structure of aspergilloglutamic peptidase from Aspergillus niger

Abstract

Aspergilloglutamic peptidase from Aspergillus niger is a novel pepstatin-insensitive acid endopeptidase distinct from the well-studied aspartic peptidases, and thus is an interesting target for protein structure/function studies. In the present study, we have determined the three-dimensional structure of the enzyme by X-ray crystallography to a 1.4-Å resolution. The results revealed that the enzyme has a unique structure, composed of two seven-stranded anti-parallel β-sheets which form a β-sandwich structure and appear to have a partial two-fold symmetry, suggesting its possible evolution by gene duplication and that the glutamic acid-110 and glutamine-24 in the heavy chain form a catalytic dyad, consistent with our results obtained by site-directed mutagenesis.

Catalytic residues and substrate specificity of scytalidoglutamic peptidase, the first member of the eqolisin in family (G1) of peptidases

“…Quite recently, the three-dimensional structure of ANCP has been elucidated [17]. Taken together with mutational analysis [18], they have proposed the same catalytic mechanism as ours [9].…”

Crystal Structure of Scytalidoglutamic Peptidase with its First Potent Inhibitor Provides Insights into Substrate Specificity and Catalysis

“…15 Furthermore, the crystal structure of AGP has also been recently determined (pdb id: 1Y43), and is found to be very similar to that of SGP. 16 From a series of fluorescence resonance energy transfer-based assays on oligopeptide substrates of SGP, a unique substrate preference particularly at the P3, P1′ and P3′ positions has been identified among those proteinases having an optimum pH in the acidic range. 14 In particular, SGP prefers a positively charged amino acid (Arg or Lys) at the P3 and P3′ positions and a smaller amino acid (Ala, Gly or Thr) at the P1′ position.…”

A proposed catalytic mechanism of aspergilloglutamic peptidase from Aspergillus niger

“…This was further confirmed by X-ray crystallographic studies, 8) which has revealed that the enzyme indeed has a unique structure, with an apparent partial two-fold symmetry, where the essential glutamic acid (Glu B110) and glutamine (Gln B24) residues identified by site-directed mutagenesis apparently form a catalytic dyad.…”

“…Figure 2a is a GRASP 15) diagram based on the crystal sructure 8) of AGP complexed by docking calculation with a dipeptide His-Ser, which represents the P1-P1' residues of a propeptide-based substrate (Lys-Arg-His--Ser-AsnAla-Ala-Tyr-Ile), indicating the electrostatic potential of the protein surface. There is a deep groove lying horizontally across the center of the molecule, which is clearly the most negative area in the protein, including the two catalytic residues, between which lies the dipeptide.…”

“…Figure 2b indicates the location of hydrophobic residues and acidic residues present within or in the vicinity of the groove based on the crystal structure. 8) It is obvious that the catalytic site and its vicinity are markedly hydrophobic; the presence of a cluster of three tryptophan residues (Trp B41, Trp B10 and Trp A7, from left to right) is especially notable. All these tryptophan residues are sitting on the bottom of the groove.…”