# Molecular Mechanisms, Pathways and Inhibition of Acetyl-Transfer Reactions

> **NIH NIH R35** · UNIVERSITY OF PENNSYLVANIA · 2022 · $232,655

## Abstract

The goal of this administrative supplement of the parent award, R35GM118090, is to purchase a TwoMP Mass
Photometer from REFEYN for the analysis of a large number of diverse macromolecular assemblies relevant
to the parent award. Many of the systems under study in the parent award mediate their biological functions
through the formation of macromolecular assemblies, and a reliable characterization of these assemblies is
essential for drawing information about biological function and for optimizing samples for subsequent structural
characterization using X-ray crystallography or single particle (SP) cryo-EM. Mass Photometry is a label-free
single molecule solution technique that will allow us to rapidly (minutes per sample), accurately, and with
relatively low sample requirement (~10 μL of a ~20 nM sample) and several-minute turnaround, analyze
various homomeric and heteromeric macromolecular assemblies ranging in size from 30 kDa – 5 MDa, yielding
information about molecular mass, distribution among oligomers, stoichiometry, and kinetics and dissociation
constants in the range of ~20 pM to ~500 nM. We will use the TwoMP Mass Photometer to analyze
macromolecular assemblies including proteins and nucleic acids under a variety of buffer and liganded
conditions. The effect of site directed mutants on macromolecular interactions will also be evaluated using
Mass Photometry. In addition, because of the exceedingly low concentration and volume of sample required
for the technology, we will be able to analyze samples from eukaryotic expression systems that are typically
produced in low yield that could not otherwise be analyzed using other biophysical approaches. These Mass
Photometry analyses will also play a key role in identifying suitable conditions for subsequent X-ray
crystallographic and SP cryo-EM analysis of macromolecules and their complexes. Specific projects that will
benefit from the instrumentation include studies on multiprotein protein and RNA acetyltransferases (Sas2/4/5,
Caf1 and Nat10), N-terminal protein acetyltransferase (NAT) regulation (NAT complexes and ribosomes), and
small molecule acetyl-transfer enzymes (ACLY, ACSS2, FASN, ACC1). There is currently no Mass
Photometer or comparable instrument on campus. The instrument will be located in a dedicated biophysical
equipment room within the Marmorstein laboratory and Dr. Marmorstein’s home Department of Biochemistry
and Biophysics is committed to the proper installation and maintenance of the Mass Photometer throughout its
lifetime.

## Key facts

- **NIH application ID:** 10581921
- **Project number:** 3R35GM118090-07S1
- **Recipient organization:** UNIVERSITY OF PENNSYLVANIA
- **Principal Investigator:** Ronen Marmorstein
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $232,655
- **Award type:** 3
- **Project period:** 2016-07-01 → 2026-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10581921

## Citation

> US National Institutes of Health, RePORTER application 10581921, Molecular Mechanisms, Pathways and Inhibition of Acetyl-Transfer Reactions (3R35GM118090-07S1). Retrieved via AI Analytics 2026-05-28 from https://api.ai-analytics.org/grant/nih/10581921. Licensed CC0.

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