Mannitol-1-phosphate dehydrogenase activity in Ectocarpus siliculosus, a key role for mannitol synthesis in brown algae.
Mannitol represents a major end product of photosynthesis in brown algae (Phaeophyceae), and is, with the β-1,3-glucan laminarin, the main form of carbon storage for these organisms. Despite its importance, little is known about the genes and enzymes responsible for the metabolism of mannitol in these seaweeds. Taking benefit of the sequencing of the Ectocarpus siliculosus genome, we focussed our attention on the first step of the synthesis of mannitol (reduction of the photo-assimilate fructose-6-phosphate), catalysed by the mannitol-1-phosphate dehydrogenase (M1PDH). This activity was measured in algal extracts, and was shown to be regulated by NaCl concentration in the reaction medium. Genomic analysis revealed the presence of three putative M1PDH genes (named EsM1PHD1, EsM1PDH2 and EsM1PDH3). Sequence comparison with orthologs demonstrates the modular architecture of EsM1PHD1 and EsM1PDH2, with an additional N-terminal domain of unknown function. In addition, gene expression experiments carried out on samples harvested through the diurnal cycle, and after several short-term saline and oxidative stress treatments, showed that EsM1PDH1 is the most highly expressed of these genes, whatever the conditions tested. In order to assess the activity of the corresponding protein, this gene was expressed in Escherichia coli. Cell-free extracts prepared from bacteria containing EsM1PDH1 displayed higher M1PDH activity than bacteria transformed with an empty plasmid. Further characterisation of recombinant EsM1PDH1 activity revealed its very narrow substrate specificity, salt regulation, and sensitivity towards an inhibitor of SH-enzymes.
Rousvoal
Sylvie
S
UPMC Univ Paris 6, UMR 7139 Marine Plants and Biomolecules, Station Biologique, 29682 Roscoff, France.
Groisillier
Agnès
A
Dittami
Simon M
SM
Michel
Gurvan
G
Boyen
Catherine
C
Tonon
Thierry
T
D000595:Amino Acid Sequence; D002940:Circadian Rhythm; D020869:Gene Expression Profiling; D018506:Gene Expression Regulation, Plant / Q000502:physiology*; D008353:Mannitol / Q000378:metabolism*; D000459:Phaeophyceae / Q000201:enzymology*; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D013312:Stress, Physiological; D013401:Sugar Alcohol Dehydrogenases / Q000235:genetics / Q000378:metabolism*
D010940:Plant Proteins; D008353:Mannitol; D013401:Sugar Alcohol Dehydrogenases; C021544:mannitol-1-phosphate dehydrogenase