Project Summary/Abstract
Ectodomain shedding is a fundamental process in biology that enables a cell to change the membrane protein
topology, affecting cell adhesion and signal transduction pathways. Accordingly, the enzymes that function in
ectodomain shedding are vital to human health and dysregulated in disease pathogenesis. Despite the
importance of these enzymes in human health, the molecular and atomic details that govern zymogen
maturation, activation, substrate selectivity, macromolecular inhibition and subcellular compartmentalization are
limited to a few examples. The long-term goal of my research program is to study the mechanistic details
of membrane tethered enzymes, revealing the universal concepts in ectodomain shedding that underpin
enzyme function and the unique properties that position an enzyme into a biological niche. Initially, we
are prioritizing the study of the a disintegrin and metalloproteinase family, specifically ADAM17, to launch my
research program as a new principal investigator.
ADAMs are complex multidomain type1 transmembrane enzyme that have essential functions in development,
fertilization and immunity. ADAM17 is credited to process over 80 different protein substrates and have essential
functions in immunity, skin morphogenesis and development of heart valve formation. All ADAMs are initially
translated as a zymogen, bound by a prodomain that functions to control enzyme latency. ADAM17 zymogen
maturation and transport through the cell is dependent on an interaction with a family of inactive rhomboid
proteins, iRhom1 and -2. Appropriately, iRhoms are positioned as vital regulators of ADAM17 activity. Despite
ADAM17s well characterized roles in development and homeostasis, significant shortcomings persist in
understanding the molecular events that regulate enzyme activity, such as zymogen maturation, enzyme
activation and association with regulatory proteins. To address these knowledge gaps, my research program will
bring new innovative approaches to the field and leverage my experience in structural and cell biology to
characterize the atomic details that regulate ADAM17 activity. Specifically, we will determine the atomic details
of the ADAM prodomain and uncover the properties that lead to enzyme specificity and inhibition. In addition,
we propose to use X-ray crystallography and electron microscopy to reveal the mechanistic details into ADAM17
activation and recognition by the accessory proteins, iRhom1 and 2. Finally, we will use an unbiased proteomic
mass spectroscopy approach to specifically evaluate the ADAM17-iRhom complex during enzyme activation in
addition to identifying the subcellular compartment and activity modulating proteins in regulating ADAM17
activity. In the next five years, we aim to develop a unified model for ADAM17 regulation and construct the
foundation to expand our efforts to include additional membrane tethered proteases and families that function in
ectodomain shedding.