1K7W

Crystal Structure of S283A Duck Delta 2 Crystallin Mutant

Summary for 1K7W

• Entry DOI: 10.2210/pdb1k7w/pdb
• Related: 1AOS 1AUW 1DCN 1HY0 1HY1 1I0A 1K62
• Descriptor: delta 2 crystallin, ARGININOSUCCINATE (3 entities in total)
• Functional Keywords: eye lens protein, delta 2 crystallin, argininosuccinate lyase, enzyme mechanism, lyase
• Biological source: Anas platyrhynchos
• Total number of polymer chains: 4
• Total formula weight: 208134.51

Authors

Sampaleanu, L.M.,Yu, B.,Howell, P.L. (deposition date: 2001-10-22, release date: 2002-03-06, Last modification date: 2024-10-02)

Primary citation

Sampaleanu, L.M.,Yu, B.,Howell, P.L.
Mutational analysis of duck delta 2 crystallin and the structure of an inactive mutant with bound substrate provide insight into the enzymatic mechanism of argininosuccinate lyase.
J.Biol.Chem., 277:4166-4175, 2002

PubMed Abstract: The major soluble avian eye lens protein, delta crystallin, is highly homologous to the housekeeping enzyme argininosuccinate lyase (ASL). ASL is part of the urea and arginine-citrulline cycles and catalyzes the reversible breakdown of argininosuccinate to arginine and fumarate. In duck lenses, there are two delta crystallin isoforms that are 94% identical in amino acid sequence. Only the delta2 isoform has maintained ASL activity and has been used to investigate the enzymatic mechanism of ASL. The role of the active site residues Ser-29, Asp-33, Asp-89, Asn-116, Thr-161, His-162, Arg-238, Thr-281, Ser-283, Asn-291, Asp-293, Glu-296, Lys-325, Asp-330, and Lys-331 have been investigated by site-directed mutagenesis, and the structure of the inactive duck delta2 crystallin (ddeltac2) mutant S283A with bound argininosuccinate was determined at 1.96 A resolution. The S283A mutation does not interfere with substrate binding, because the 280's loop (residues 270-290) is in the open conformation and Ala-283 is more than 7 A from the substrate. The substrate is bound in a different conformation to that observed previously indicating a large degree of conformational flexibility in the fumarate moiety when the 280's loop is in the open conformation. The structure of the S283A ddeltac2 mutant and mutagenesis results reveal that a complex network of interactions of both protein residues and water molecules are involved in substrate binding and specificity. Small changes even to residues not involved directly in anchoring the argininosuccinate have a significant effect on catalysis. The results suggest that either His-162 or Thr-161 are responsible for proton abstraction and reinforce the putative role of Ser-283 as the catalytic acid, although we cannot eliminate the possibility that arginine is released in an uncharged form, with the solvent providing the required proton. A detailed enzymatic mechanism of ASL/ddeltac2 is presented.

PubMed: 11698398
DOI: 10.1074/jbc.M107465200

Experimental method

X-RAY DIFFRACTION (1.96 Å)