Identification and characterization of a unique cysteine residue proximal to the catalytic site of Arabidopsis thaliana carotenoid cleavage enzyme 1

Download
  1. Get@NRC: Identification and characterization of a unique cysteine residue proximal to the catalytic site of Arabidopsis thaliana carotenoid cleavage enzyme 1 (Opens in a new window)
DOIResolve DOI: http://doi.org/10.1139/O08-031
AuthorSearch for: ; Search for: ; Search for: ; Search for:
TypeArticle
Journal titleBiochemistry and Cell Biology
Volume86
Issue3
Pages262270; # of pages: 9
Subjectcarotenoid cleavage enzymes; carotenoids; type II Cu center; cysteine mutagenesis
AbstractAtCCD1 and AtNCED3 are related carotenoid cleavage enzymes from Arabidopsis thaliana that catalyze the oxidative cleavage of, respectively, the 9,10 (9′,10′) double bonds of carotenoid substrates such as β-carotene, and the 11,12 double bond of 9-cis epoxycarotenoids. Although the cellular and cleavage functionalities of these enzymes have been reported, their mechanisms and related structural environments mediating these disparate specificities in homologous enzymes have not been well characterized. By relating the differences observed in UV and visible light absorption and Cu(II) electron paramagnetic signals to variations in sequence alignments and 3-D homology models of the two A. thaliana enzymes, we identified a putatively proximal cysteine residue (Cys352) in AtCCD1 that is not conserved in AtNCED3. Spectral analysis of the Cys to Ala mutant confirmed its uniqueness and proximity to the metal binding site, but precluded any role for the residue in the mediation of the observed metal binding affinity or associated steric constraint differences. Further analysis of kinetic substrate cleavage properties indicated a decrease in Vmax and a subtle increase in Km for the C352A mutant compared with those observed for the wild-type, thus confirming catalytic site proximity and suggesting possible roles for the unique cysteine in the modulation of substrate affinity and (or) the reaction rate of AtCCD1.
Publication date
LanguageEnglish
AffiliationNational Research Council Canada; NRC Plant Biotechnology Institute
Peer reviewedNo
NPARC number21187430
Export citationExport as RIS
Report a correctionReport a correction
Record identifier5f2c2209-073a-43e6-a8dc-3460aa8aa303
Record created2013-01-10
Record modified2016-05-09
Bookmark and share
  • Share this page with Facebook (Opens in a new window)
  • Share this page with Twitter (Opens in a new window)
  • Share this page with Google+ (Opens in a new window)
  • Share this page with Delicious (Opens in a new window)