Niels Bradshaw, Ph.D.
Assistant Professor
Department of Biochemistry
Brandeis University
October 1, 2018
Regulation of Protein Phosphatases and the Evolution of Cellular Signaling
Phosphorylation can provide reversible functional regulation of proteins in a cell. This process depends on kinases and phosphatases, though their role is far from well-understood. Dr. Bradshaw and his lab focus on particular phosphatases, PP2C SpollE, and how they are regulated. His work has determined that PP2C phosphatases are specific to certain substrates and are regulated themselves by a biochemical switch.
Reversible phosphorylation is an important and widespread biological regulatory mechanism. Kinases and phosphatases integrate diverse cellular and environmental cues to precisely tune the activity of target proteins by controlling their phosphorylation state. Despite early appreciation that phosphatases are the direct targets of regulatory inputs and are highly substrate-specific, the roles of phosphatases as regulatory elements in signaling systems have largely been overlooked. As a result, how phosphatases are regulated and directed to specific protein substrates is a major outstanding question.
PP2C serine/threonine phosphatases control biological processes including stress responses, development, and cell division in all kingdoms of life. Diverse cis-regulatory domains adapt PP2C phosphatases to specific functions, but how these domains control phosphatase activity was unknown. Dr. Bradshaw determined structures representing active and inactive states of the PP2C phosphatase SpoIIE from Bacillus subtilis. Based on structural analysis and genetic and biochemical experiments, he identified an α-helical switch that shifts a carbonyl oxygen into the active site to coordinate a metal cofactor. This switch is widely conserved among PP2C family members, coupling phosphatase activity to diverse inputs. This provides a potential mechanism for how PP2C phosphatases are subject to regulation by diverse regulatory domains that respond to varied inputs.
Using paralogous signaling systems that exist in the same cell, Dr. Bradshaw found that PP2C phosphatases are highly specific for their cognate substrate proteins and that this specificity is achieved through preferential substrate binding, catalysis, and cofactor binding for cognate substrates. The switch is significantly responsible for this selectivity, ensures that phosphatase is only active when bound to the appropriate substrate and thus acts as a unified platform for regulation and substrate specificity.
Remarkably, Dr. Bradshaw found that the switch is shared with proteasomal proteases, which he identified as evolutionary and structural relatives of PP2C phosphatases. Although these proteases use an unrelated catalytic mechanism, rotation of equivalent helices controls protease activity by movement of the equivalent carbonyl oxygen into the active site. This suggests that these two divergent families of signaling proteins may be ancient evolutionary relatives linked by a common structural fold and regulatory mechanism.