# Evolution of effector caspase conformational landscapes

> **NIH NIH R01** · UNIVERSITY OF TEXAS ARLINGTON · 2021 · $293,296

## Abstract

Caspases are an ancient class of cysteinyl proteases that are critical to apoptosis and cell
differentiation, but little is known about how caspase activity is modulated for apoptotic versus non-
apoptotic events. Our primary goal in this proposal is to determine evolutionary trajectories from a
common scaffold that resulted in modulation of activity in extant caspase clusters. The apoptotic caspases
evolved from a common ancestor into two distinct subfamilies that are either monomers (initiator
caspases) or dimers (effector caspases). Biologically, dimerization of initiator caspases is an important
cell-fate determining property; conversely, dimeric effector caspases evolved unique allosteric
mechanisms to fine-tune activity. Our long-term goal is to integrate evolutionary biology with rigorous
biochemical studies to define the evolutionary trajectories in caspases. These studies will stimulate new
areas for evolutionary biochemical studies on apoptotic regulatory mechanisms, protein oligomerization,
and developing enzymes with altered specificities. The objective of this grant is to characterize mutations
that occurred in the common ancestor that resulted in two distinct subfamilies of apoptotic caspases. The
central hypothesis is that limited mutations in the caspase-hemoglobinase scaffold established the folding
and conformational landscapes >650 million years ago and that extant caspases evolved different
properties through differentially modifying common interaction networks. Our rationale is that studies of
reconstructed ancestral proteins suggest that mutations in a weak ancestral dimer resulted in two
subfamilies with different oligomeric properties. Neofunctionalization of the two subfamilies provided
distinct enzyme substrate selection as well as allosteric mechanisms to modulate activity. Our specific
aims will test the following hypotheses: (Aim1) A weakly dimeric common ancestor provided a platform for
evolution of dimeric and monomeric caspase subfamilies; (Aim 2) A promiscuous common ancestor
provided a platform for minimal modifications that resulted in modern enzyme selection; (Aim 3)
Evolutionary changes in a common allosteric network resulted in unique regulatory mechanisms through
selection of different conformations in the native ensemble. This contribution is significant since it will
establish key features of substrate specificity and allosteric regulation that were retained through hundreds
of millions of years of evolution, while other regulatory features are modern and cluster-specific. The
proposed research is innovative because we established a database (CaspBase) containing over 6,600
caspase sequences from 353 taxa in order to infer ancestral sequences and reconstruct ancestral
proteins, which are used to characterize evolutionary trajectories. Insight into caspase evolution is
impactful because properties retained for more than 700 million years in the ancestral scaffold can be
used to generate new enzymes with alt...

## Key facts

- **NIH application ID:** 10130380
- **Project number:** 5R01GM127654-03
- **Recipient organization:** UNIVERSITY OF TEXAS ARLINGTON
- **Principal Investigator:** ALLAN CLAY CLARK
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $293,296
- **Award type:** 5
- **Project period:** 2019-04-01 → 2023-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10130380

## Citation

> US National Institutes of Health, RePORTER application 10130380, Evolution of effector caspase conformational landscapes (5R01GM127654-03). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10130380. Licensed CC0.

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