# Structure and dynamics of clinically-relevant cytochrome P450 enzymes

> **NIH NIH R01** · UNIVERSITY OF MICHIGAN AT ANN ARBOR · 2020 · $429,111

## Abstract

ABSTRACT
Human cytochrome P450 enzymes are dynamic, often promiscuous, monooxygenases. Some function in the
biosynthesis of critical endogenous compounds and are frequent drug targets. Others are dominant factors in
drug metabolism, dictating drug clearance and/or prodrug activation. For both, understanding P450 interactions
with substrates, inhibitors, and their catalytic partner proteins provides substantial useful information in drug
design. While X-ray structures have provided numerous insights into drug binding to key human P450 enzymes,
there are many gaps that cannot be filled by this approach. Conformational changes that P450s must undergo
to channel ligands to the active site and for a single P450 to accommodate many different small molecule
scaffolds only randomly become apparent in comparing the X-ray structures achievable. Despite substantial
efforts, some key human P450 enzymes have not yielded to crystallization. Many drug substrates have lower
active site affinity and/or multiple orientations not suitable to determining clear X-ray structures. There are no
structures of human P450 enzymes with reductase or cytochrome b5 catalytic partners. As a result, drug design
is limited by available structural information.
We propose employing solution NMR as a newly-viable orthogonal method to obtain the requisite amino acid-
level structural information needed to understand human P450/ligand and P450/protein interactions. While the
size, stability, and the absence of information relating individual NMR resonances to the corresponding amino
acid have all thus far prevented the determination of any human P450 structures by NMR, we have the combined
expertise and preliminary data to demonstrate that this feat is now technically possible. The Pochapsky lab
previously developed the expertise to determine solution NMR structures of slightly smaller, soluble bacterial
P450 enzymes. The Scott lab developed the capacity to generate human membrane P450 enzymes in the
isotopically-labeled forms, with the amounts and with the stability required for NMR experiments. Thus, based
on substantial preliminary data, we propose to 1) advance strategies for determining human P450 structures by
solution NMR while determining the human steroidogenic CYP17A1 structure and 2) apply solution NMR
strategies to the two most important human drug-metabolizing P450 enzymes, CYP3A4 and CYP2D6.
Successful completion of aim 1 will not only further establish the feasibility of NMR structures for human
membrane P450 enzymes, but will do so for an important prostate cancer drug target for which additional
structural information is essential to further drug design. Results of both aims will provide a reference set of
amino acid assignments that can then be readily used by a wide range of non-NMR experts, in much simpler
experiments, to quickly determine where and how drugs and other proteins bind to three clinically-important
P450 enzymes, as well as deciphering the changes in p...

## Key facts

- **NIH application ID:** 9842541
- **Project number:** 5R01GM130997-02
- **Recipient organization:** UNIVERSITY OF MICHIGAN AT ANN ARBOR
- **Principal Investigator:** Thomas Charles Pochapsky
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $429,111
- **Award type:** 5
- **Project period:** 2019-01-01 → 2022-11-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9842541

## Citation

> US National Institutes of Health, RePORTER application 9842541, Structure and dynamics of clinically-relevant cytochrome P450 enzymes (5R01GM130997-02). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/9842541. Licensed CC0.

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