PDBsum entry 1bcr

Hydrolase/hydrolase inhibitor

PDB id

1bcr

Contents

			Protein chains

					254 a.a. *


					152 a.a. *

			Ligands

					VAL-OAR


					NAG-FUC-NAG


					NAG-NAG


					NAG


					ARG

			Waters ×192

* Residue conservation analysis

References listed in PDB file

Key reference

Title

Peptide aldehyde complexes with wheat serine carboxypeptidase ii: implications for the catalytic mechanism and substrate specificity.

Authors

T.L.Bullock, K.Breddam, S.J.Remington.

Ref.

J Mol Biol, 1996, 255, 714-725. [DOI no: 10.1006/jmbi.1996.0058]

PubMed id

8636973

Abstract

The structures of two ternary complexes of wheat serine carboxypeptidase II (CPD-WII), with a tetrapeptide aldehyde and a reaction product arginine, have been determined by X-ray crystallography at room temperature and -170 degrees. The peptide aldehydes, antipain and chymostatin, form covalent adducts with the active-site serine 146. The CPD-WII antipain arginine model has a standard crystallographic R-factor of 0.162, with good geometry at 2.5 A resolution for data collected at room temperature. The -170 degrees C model of the chymostatin arginine complex has an R-factor of 0.174, with good geometry using data to 2.1 A resolution. The structures suggest binding subsites N-terminal to the scissile bond. All four residues of chymostatin are well-localized in the putative S1 through S4 sites, while density is apparent only in S1 and S2 for antipain. In the S1 site, Val340 and 341, Phe215 and Leu216 form a hydrophobic binding surface, not a pocket, for the P1 phenylalanyl side-chain of chymostatin. The P1 arginyl of antipain also binds at this site, but the positive charge appears to be stabilized by additional solvent molecules. Thus, the hybrid nature of the S1 site accounts for the ability of CPD-WII to accept both hydrophobic and basic residues at P1. Hydrogen bonds to the peptide substrate backbone are few and are made primarily with side-chains on the enzyme. Thus, substrate recognition by CPD-WII appears to have nothing in common with that of the other families of serine proteinases. The hemiacetal linkages to the essential Ser146 are of a single stereoisomer with tetrahedral geometry, with an oxygen atom occupying the "oxyanion hole" region of the enzyme. This atom accepts three hydrogen bonds, two from the polypeptide backbone and one from the positively-charged amino group of bound arginine, and must be negatively charged. Thus, the combination of ligands forms an excellent approximation to the oxyanion intermediate formed during peptide hydrolysis. Surprisingly, the (R) stereochemistry at the hemiacetal linkage is opposite to that expected by comparison to previously determined structures of peptide aldehydes complexed with Streptomyces griseus proteinase A. This is shown to be a consequence of the approximate mirror symmetry of the arrangement of catalytic groups in the two families of serine proteases and suggests that the stereochemical course of the two enzymatic reactions differ in handedness.



	Figure 1. Figure 1. The chemical structures of the peptide aldehydes, antipain and chymostatin from Streptomyces. Chymostatin is isolated as a mixture of three forms, A, B, and C. These have R groups of leucine (A), valine (B), or isoleucine (C) side-chains.		Figure 5. Figure 5. (a) Stereo view of the hydrogen bonding pattern for chy- mostatin (filled bonds) from the refined -170° model. The hydrogen bonds are shown as fine lines and have the indicated lengths in Å . The adjoining residues (open bonds) of CPD-WII form one wall of the active site cavity. (b) Schematic diagram of the hydrogen bond and van der Waals' interactions of the chymo- statin/CPD-WII complex. The bold curves indicate van der Waals' contacts with side-chains of the residues indicated, while broken (b) lines indicate hydrogen bonds.