TY - JOUR
T1 - Comprehensive survey of redox sensitive starch metabolising enzymes in Arabidopsis thaliana
AU - Glaring, Mikkel Andreas
AU - Skryhan, Katsiaryna
AU - Kötting, Oliver
AU - Zeeman, Samuel C.
AU - Blennow, Per Gunnar Andreas
PY - 2012/9
Y1 - 2012/9
N2 - In chloroplasts, the ferredoxin/thioredoxin pathway regulates enzyme activity in response to light by reduction of regulatory disulfides in target enzymes, ensuring coordination between photosynthesis and diurnal metabolism. Although earlier studies have suggested that many starch metabolic enzymes are similarly regulated, redox regulation has only been verified for a few of these in vitro. Using zymograms and enzyme assays, we performed a comprehensive analysis of the redox sensitivity of known starch metabolising enzymes in extracts of . Arabidopsis thaliana. Manipulation of redox potentials revealed that several enzymatic activities where activated by reduction at physiologically relevant potentials. Among these where the isoamylase complex . AtISA1/. AtISA2, the limit dextrinase . AtLDA, starch synthases . AtSS1 and . AtSS3, and the starch branching enzyme . AtBE2. The reversibility of the redox reaction was confirmed by enzyme assays for . AtLDA, . AtSS1 and . AtSS3. Analysis of an . AtBAM1 knock-out mutant identified an additional redox sensitive β-amylase activity, which was most likely . AtBAM3. A similar requirement for reducing conditions was observed for recombinant chloroplastic α-amylase (. AtAMY3) activity. This study adds further candidates to the list of reductively activated starch metabolising enzymes and supports the view that redox regulation plays a role in starch metabolism.
AB - In chloroplasts, the ferredoxin/thioredoxin pathway regulates enzyme activity in response to light by reduction of regulatory disulfides in target enzymes, ensuring coordination between photosynthesis and diurnal metabolism. Although earlier studies have suggested that many starch metabolic enzymes are similarly regulated, redox regulation has only been verified for a few of these in vitro. Using zymograms and enzyme assays, we performed a comprehensive analysis of the redox sensitivity of known starch metabolising enzymes in extracts of . Arabidopsis thaliana. Manipulation of redox potentials revealed that several enzymatic activities where activated by reduction at physiologically relevant potentials. Among these where the isoamylase complex . AtISA1/. AtISA2, the limit dextrinase . AtLDA, starch synthases . AtSS1 and . AtSS3, and the starch branching enzyme . AtBE2. The reversibility of the redox reaction was confirmed by enzyme assays for . AtLDA, . AtSS1 and . AtSS3. Analysis of an . AtBAM1 knock-out mutant identified an additional redox sensitive β-amylase activity, which was most likely . AtBAM3. A similar requirement for reducing conditions was observed for recombinant chloroplastic α-amylase (. AtAMY3) activity. This study adds further candidates to the list of reductively activated starch metabolising enzymes and supports the view that redox regulation plays a role in starch metabolism.
U2 - 10.1016/j.plaphy.2012.06.017
DO - 10.1016/j.plaphy.2012.06.017
M3 - Journal article
SN - 0981-9428
VL - 58
SP - 89
EP - 97
JO - Plant Physiology and Biochemistry
JF - Plant Physiology and Biochemistry
ER -