# Molecular characterization of cardiomyopathy mutations in human cardiac myosin

> **NIH NIH R01** · UNIVERSITY OF COLORADO · 2020 · $583,148

## Abstract

PROJECT SUMMARY / ABSTRACT
The 10-member sarcomeric myosin heavy chain gene family has been studied extensively and mutations in
half of its members have been implicated in disease. Myosin motors convert chemical energy into mechanical
force by amplifying the ATP-driven conformational rotation of myosin’s lever arm, which consists of a helical
pliant region and the α-helix of the heavy chain stabilized and stiffened by essential and regulatory light
chains; hereafter referred to as the lever arm. While a great deal of study has been devoted to the catalytic
domain and converter, the lever arm has often been treated as simply a semi-rigid extension of the converter
to amplify the stroke size of the motor. For a given isoform of myosin the entire lever arm is highly conserved
across species, but it is highly variant amongst the 10 isoforms, suggesting that the sequences of lever arm α-
helices confer specific functions. Given the high sequence conservation of the β-cardiac lever arm across
species and the high density of pathogenic mutations in it, we hypothesize that this region is an important
regulatory domain that modulates myosin function and testing that hypothesis is the focus of this proposal.
We propose an interdisciplinary collaboration among the Spudich, Perkins and Leinwand laboratories to study
the effects of disease-causing mutations in the lever arm by integrating the biophysical characterization of
isolated lever arms and myosin motor functional assays with cardiac cell biology. In the previous grant period,
the Leinwand, Spudich and Geeves laboratories produced and characterized a number of disease-causing
mutations of the human β-cardiac myosin motor for their biochemical and kinetic properties, but none of these
studies included lever arm mutations. Because of clinical hypercontractility of hypertrophic cardiomyopathy
(HCM) patients and hypocontractility of dilated cardiomyopathy (DCM) patients, we hypothesize that HCM
mutations will most likely increase the stiffness of the lever arm, whereas, DCM mutations will cause the lever
arm to be less stiff. In Aim I, we will determine the biochemical and mechanical properties of the lever arm of
WT β-myosin using atomic force microscopy (AFM). AFM has emerged as a powerful tool for investigating
the elasticity of proteins in addition to probing their folding/unfolding dynamics. Until now, AFM technology did
not have the resolution to study the mechanics of the 9-nm long lever arm. However, the Perkins lab’s recent
advances in single-molecule AFM techniques will enable us to compare the mechanical properties of the WT
ß-cardiac myosin lever arm α-helix to ones carrying cardiomyopathy-causing mutations. In Aim II, we will
measure the impact of the lever arm mutations on in vitro subfragment-1 (S1) motor function using ATPase,
gliding filament and optical tweezer assays. Finally, in Aim III we will integrate these biophysical and
biomechanical findings into cells by introducing WT and leve...

## Key facts

- **NIH application ID:** 9847984
- **Project number:** 5R01HL117138-07
- **Recipient organization:** UNIVERSITY OF COLORADO
- **Principal Investigator:** Leslie Anne Leinwand
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $583,148
- **Award type:** 5
- **Project period:** 2013-08-20 → 2022-01-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9847984

## Citation

> US National Institutes of Health, RePORTER application 9847984, Molecular characterization of cardiomyopathy mutations in human cardiac myosin (5R01HL117138-07). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/9847984. Licensed CC0.

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