Oligomerization and carbohydrate binding of glucan phosphatases

Glucan phosphatases are a unique subset of the phosphatase family that bind to and dephosphorylate carbohydrate substrates. Family members are found in diverse organisms ranging from single-cell red algae to humans. The nature of their functional oligomerization has been a source of considerable debate. We demonstrate that the human laforin protein behaves aberrantly when subjected to Size Exclusion Chromotography (SEC) analysis due to interaction with the carbohydrate-based matrix. This interaction complicates the analysis of laforin human disease mutations. Herein, we show that SEC with Multi-Angle static Light Scattering (SEC-MALS) provides a method to robustly define the oligomerization state of laforin and laforin variants. We further analyzed glucan phosphatases from photosynthetic organisms to define if this interaction was characteristic of all glucan phosphatases. Starch EXcess-four (SEX4) from green plants was found to lack significant interaction with the matrix and instead exists as a monomer. Conversely, Cm-laforin, from red algae, exists as a monomer in solution while still exhibiting significant interaction with the matrix. These data demonstrate a range of oligomerization behaviors of members of the glucan phosphatase family, and establish SEC-MALS as a robust methodology to quantify and compare oligomerization states between different proteins and protein variants in this family.