δ Crystallin, a taxon-specific crystallin present in avian eye lenses, is homologous to the urea cycle enzyme ASL (argininosuccinate lyase). Although there are two δ crystallin isoforms in duck lenses, dδc1 (duck δ1 crystallin) and dδc2 (duck δ2 crystallin), only dδc2 is catalytically active. Previous structural studies have suggested that residues Ser283 and His162 in the multi-subunit active site of dδc2/ASL are the putative catalytic acid/base, while the highly conserved, positively charged Lys289 is thought to help stabilize the carbanion intermediate. The strict conservation of a small hydroxy-containing residue (Thr or Ser) at position 161 adjacent to the putative catalytic base, as well as its proximity to the substrate in the S283A dδc2 enzyme–substrate complex, prompted us to investigate further the role this residue. Structures of the active T161S and inactive T161D dδc2 mutants, as well as T161D complexed with argininosuccinate, have been determined to 2.0 Å resolution. The structures suggest that a hydroxy group is required at position 161 to help correctly position the side chain of Lys289 and the fumarate moiety of the substrate. Threonine is probably favoured over serine, because the interaction of its methyl group with Leu206 would restrict its conformational flexibility. Residues larger than Thr or Ser interfere with substrate binding, supporting previous suggestions that correct positioning of the substrate's fumarate moiety is essential for catalysis to occur. The presence of the 280s loop (i.e. a loop formed by residues 270–290) in the ‘open’ conformation suggests that loop closure, thought to be essential for sequestration of the substrate, may be triggered by the formation of the carbanion or aci-carboxylate intermediates, whose charge distribution more closely mimics that of the sulphate ion found in the active-site region of the inactive dδc1. The 280s loop in dδc1 is in the closed conformation.
Skip Nav Destination
Article navigation
December 2004
-
Cover Image
Cover Image
- PDF Icon PDF LinkFront Matter
- PDF Icon PDF LinkTable of Contents
- PDF Icon PDF LinkEditorial Board
Research Article|
November 23 2004
Structural studies of duck δ2 crystallin mutants provide insight into the role of Thr161 and the 280s loop in catalysis
Liliana M. SAMPALEANU;
Liliana M. SAMPALEANU
1
*Structural Biology and Biochemistry, Hospital for Sick Children, 555 University Avenue, Toronto, Ontario, Canada M5G 1X8
†Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada M5S 1A8
Search for other works by this author on:
Penelope W. CODDING;
Penelope W. CODDING
1
‡Chemistry Department, University of Victoria, Victoria, British Columbia, Canada V8W 3V6
Search for other works by this author on:
Yuri D. LOBSANOV;
Yuri D. LOBSANOV
*Structural Biology and Biochemistry, Hospital for Sick Children, 555 University Avenue, Toronto, Ontario, Canada M5G 1X8
Search for other works by this author on:
May TSAI;
May TSAI
*Structural Biology and Biochemistry, Hospital for Sick Children, 555 University Avenue, Toronto, Ontario, Canada M5G 1X8
†Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada M5S 1A8
Search for other works by this author on:
G. David SMITH;
G. David SMITH
*Structural Biology and Biochemistry, Hospital for Sick Children, 555 University Avenue, Toronto, Ontario, Canada M5G 1X8
Search for other works by this author on:
Cathy HORVATIN;
Cathy HORVATIN
*Structural Biology and Biochemistry, Hospital for Sick Children, 555 University Avenue, Toronto, Ontario, Canada M5G 1X8
Search for other works by this author on:
P. Lynne HOWELL
P. Lynne HOWELL
2
*Structural Biology and Biochemistry, Hospital for Sick Children, 555 University Avenue, Toronto, Ontario, Canada M5G 1X8
†Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada M5S 1A8
2To whom correspondence should be addressed (email howell@sickkids.ca).
Search for other works by this author on:
Publisher: Portland Press Ltd
Received:
April 20 2004
Revision Received:
June 22 2004
Accepted:
August 23 2004
Accepted Manuscript online:
August 23 2004
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London
2004
Biochem J (2004) 384 (2): 437–447.
Article history
Received:
April 20 2004
Revision Received:
June 22 2004
Accepted:
August 23 2004
Accepted Manuscript online:
August 23 2004
Citation
Liliana M. SAMPALEANU, Penelope W. CODDING, Yuri D. LOBSANOV, May TSAI, G. David SMITH, Cathy HORVATIN, P. Lynne HOWELL; Structural studies of duck δ2 crystallin mutants provide insight into the role of Thr161 and the 280s loop in catalysis. Biochem J 1 December 2004; 384 (2): 437–447. doi: https://doi.org/10.1042/BJ20040656
Download citation file:
Sign in
Don't already have an account? Register
Sign in to your personal account
You could not be signed in. Please check your email address / username and password and try again.
Captcha Validation Error. Please try again.