# Chemical Approaches to Cell Signaling Enzymes

> **NIH NIH R01** · BRIGHAM AND WOMEN'S HOSPITAL · 2020 · $425,062

## Abstract

Abstract:
This longstanding competing renewal application CA74305 concerns our plans to apply
chemical and biochemical approaches in cell signaling to enhance our understanding of
protein post-translational modifications (PTMs) and the pathways involved. In the next
cycle, we plan to investigate the regulation of three key enzymes, PTEN, WWP2, and
AKT1 that are connected to the cancer promoting phospholipid, PIP3 (phosphatidyl
3,4,5-triphosphate). PTEN is one of the most commonly mutated tumor suppressor
genes and catalyzes the conversion of PIP3 to PIP2 and in this way antagonizes the
action of the PI3-kinase oncogenic signaling pathway. WWP2 is a HECT domain E3
ligase (ubiquitin transferase) that targets PTEN for destruction and is considered to be
an oncogene. AKT1, also an oncogene, is a protein Ser/Thr kinase that is activated by
PIP3 and drives tumor formation and is an intensively studied cancer therapeutic target.
PTEN, WWP2, and AKT1 are each subjected to multiple phosphorylations that are
proposed to have important regulatory implications. Our research program will continue
to adapt and employ protein semisynthetic methods including the newly described
enzyme-catalyzed expressed protein ligation (EPL) to generate these signaling enzymes
containing the requisite PTMs and appropriate mimics to interrogate how these
phosphorylations influence the structure and function of these key cancer-related
enzymes. Our three specific aims are: 1. Optimize enzyme-catalyzed expressed protein
ligation (EPL) for protein semisynthesis and apply to the study of PTEN regulation; 2.
Elucidate the molecular basis of WWP2 E3 ubiquitin ligase regulation; 3. Determine the
role of C-terminal phosphorylation of AKT1 on its structure, activity, mechanism of
regulation, substrate specificity, and inhibitor sensitivity. With site-specifically modified
signaling enzymes in hand, we will integrate kinetic analysis, X-ray crystallography,
protein microarrays, and cell-based studies to clarify key regulatory and signaling
features. Upon completion of this research effort, we should have a much clearer
understanding of the PIP3 signaling network. Moreover, these studies should pave the
way for new therapeutic strategies to combat PTEN-AKT1 pathway dysregulation in
cancer.

## Key facts

- **NIH application ID:** 9928025
- **Project number:** 5R01CA074305-26
- **Recipient organization:** BRIGHAM AND WOMEN'S HOSPITAL
- **Principal Investigator:** PHILIP A COLE
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $425,062
- **Award type:** 5
- **Project period:** 1997-04-01 → 2022-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9928025

## Citation

> US National Institutes of Health, RePORTER application 9928025, Chemical Approaches to Cell Signaling Enzymes (5R01CA074305-26). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/9928025. Licensed CC0.

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