UCSF

This thesis examines multiple facets of the mitochondrial phosphatase Phosphoglycerate Mutase Family Member 5 (PGAM5), including its structure, function and mechanism of activation. The Phosphoglycerate mutase family, titled for its namesake phosphoglycerate Mutase (PGAM), is a family of enzymes able to catalyze the transfer of a phosphate group from 3-phosphoglycerate to 2-phosphoglycerate. Despite a conserved catalytic core and its classification into the PGAM family, PGAM5 lacks any detectable phosphoglycerate mutase activity and paradoxically possesses phosphatase activity.

PGAM5 is a mitochondrially localized serine, threonine and histidine phosphatase that is responsible for dephosphorylation of a wide variety of cytosolic protein substrates and has been identified as an interacting partner of multiple mitochondrial proteins. This mitochondrially-tethered protein has been implicated in progression of apoptosis, necrosis and mitophagy where it has been shown to promote cell death in response to oxidative stress or mitochondrial damage. In addition to its role in contributing to mitochondrial destruction and cell death, PGAM5 has been implicated in the regulation of mitochondrial fission. PGAM5’s exact switch from a possible mitochondrial remodeler to a harbinger of death is still unknown, though preliminary data suggests it may be tied to its cleavage and subsequent migration from the inner mitochondrial membrane (IMM) space to the cytosol.

When we embarked on this project in the Jura lab, the mechanism for regulation of PGAM5’s phosphatase activity was poorly understood, and as a result, the tools for understanding the biological functions of PGAM5 or manipulating its activity did not exist. Using a wide variety of techniques, among these X-ray crystallography, electron cryo-microscopy, super-resolution microscopy and activity assays, we have observed PGAM5 is able to form higher order filaments in solution and in cells and succeeded in dissecting the dodecamer that comprises these rings and leads to PGAM5’s activation as a phosphatase.

The first chapter of this thesis contains a brief introduction to PGAM5, including background information on the histidine phosphatase superfamily, of which the PGAM family of proteins is classified, the PGAM family itself and PGAM5’s diverse roles in the cell. The second chapter of this thesis focuses on exploring the way in which PGAM5’s phosphatase activity is regulated. In this chapter, we explore PGAM5’s ability to form higher order structures, dissect these structures and unravel PGAM5’s activation as a phosphatase. The third chapter focuses on our work to employ our newfound structural knowledge of PGAM5 in an effort to identify, test and optimize novel PGAM5 activators and inhibitors.