Glucan phosphatases in unicellular green algae

CARRILLO, JB; MORALES, LL; GOMEZ-CASATI, DF; BUSI, MV; MARTÍN, M

Simposio; Simposio de Genomica Funcional de Plantas; 2017

Phosphatases are enzymes that remove phosphate groupsfrom a wide variety of substrates that includes proteins, glucans, nucleicacids and lipids. Over the past decade, phosphoglucan phosphatases havesurfaced as fundamental proteins because they are essential for normal starchdegradation in plants as well as glycogen metabolism in mammals. Although manystudies have postulated that glucan phosphatase activity is required toregulate the metabolism of storage carbohydrates or amylopectin-like materialacross multiple kingdoms, there is no information available regarding theseenzymes function in green algae.  In this sense, we explored the presence and functionof glucan phosphatases in the picoalgae Ostreococcus tauri, the smallestknown free-living eukaryote, and in Chlamydomonasreinhardtii, a well-studied biological model organism. To this end,the protein sequences of starch excess 4 (SEX4) and Like SEX4 2 (LSF2) from Arabidopsisthaliana were used for a BLAST search in O. tauri and C. reinharditii transcriptome.This analysis revealed two lociencoding proteins with similarity to LSF2 in O. tauri and one locus homologous to SEX4 and another to LSF2 in C.reinharditii. To further characterize these findings, the correspondingcDNA were cloned to express and purify the recombinant proteins in E. coli cells. We verified phosphataseactivity in vitro with pNPP as well as their natural substrateamylopectin, but with a different kinetic behavior. The results foundcontribute to affirm that the green lineage of glucan phosphatases is well conserved from landplants to unicellular green algae, evidencing the essential nature ofreversible phosphorylation of glucans in green plants.