Glucan phosphatases in unicelluar green algae

GOMEZ-CASATI DF; MORALES L; MARTIN M; CARRILLO J B; BUSI MV

Rosario

Simposio; Simposio de Genómica Funcional en Plantas; 2017

Phosphatases are enzymes that remove phosphate groups from a wide variety of substrates that includes proteins, glucans, nucleic acids and lipids. Over the past decade, phosphoglucan phosphatases have surfaced as fundamental proteins because they are essential for normal starch degradation in plants as well as glycogen metabolism in mammals. Although many studies have postulated that glucan phosphatase activity is required to regulate storage carbohydrates or amylopectin-like material metabolism across multiple kingdoms, there is no information available regarding these enzymes function in green algae.In this sense, we explored the presence and function of glucan phosphatases in the picoalgae Ostreococcus tauri, the smallest known free-living eukaryote, and in Chlamydomonas reinhardtii, a well-studied biological model organism. To this end, the protein sequences of Starch Excess 4 (SEX4) and Like SEX4 2 (LSF2) from Arabidopsis thaliana were used for a BLAST search in O. tauri and C. reinharditii transcriptome. This analysis revealed two loci encoding 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 corresponding cDNA were cloned to express and purify the recombinant proteins in E. coli cells. We verified phosphatase activity in vitro with pNPP as well as their natural substrate amylopectin, but with a different kinetic behavior. Theresults found contribute to affirm that the green lineage of glucan phosphatases is well conserved from land plants to unicellular green algae, evidencing the essential nature of reversible phosphorylation of glucans in green plants.