Mechanical unfolding of cardiac myosin binding protein-C by atomic force microscopy

Árpád Karsai, Miklós S Z Kellermayer, Samantha - Harris

Research output: Contribution to journalArticle

23 Citations (Scopus)

Abstract

Cardiac myosin-binding protein-C (cMyBP-C) is a thick-filament-associated protein that performs regulatory and structural roles within cardiac sarcomeres. It is a member of the immunoglobulin (Ig) superfamily of proteins consisting of eight Ig- and three fibronectin (FNIII)-like domains, along with a unique regulatory sequence referred to as the M-domain, whose structure is unknown. Domains near the C-terminus of cMyBP-C bind tightly to myosin and mediate the association of cMyBP-C with thick (myosin-containing) filaments, whereas N-terminal domains, including the regulatory M-domain, bind reversibly to myosin S2 and/or actin. The ability of MyBP-C to bind to both myosin and actin raises the possibility that cMyBP-C cross-links myosin molecules within the thick filament and/or cross-links myosin and thin (actin-containing) filaments together. In either scenario, cMyBP-C could be under mechanical strain. However, the physical properties of cMyBP-C and its behavior under load are completely unknown. Here, we investigated the mechanical properties of recombinant baculovirus-expressed cMyBP-C using atomic force microscopy to assess the stability of individual cMyBP-C molecules in response to stretch. Force-extension curves showed the presence of long extensible segment(s) that became stretched before the unfolding of individual Ig and FNIII domains, which were evident as sawtooth peaks in force spectra. The forces required to unfold the Ig/FNIII domains at a stretch rate of 500 nm/s increased monotonically from ∼30 to ∼150 pN, suggesting a mechanical hierarchy among the different Ig/FNIII domains. Additional experiments using smaller recombinant proteins showed that the regulatory M-domain lacks significant secondary or tertiary structure and is likely an intrinsically disordered region of cMyBP-C. Together, these data indicate that cMyBP-C exhibits complex mechanical behavior under load and contains multiple domains with distinct mechanical properties.

Original languageEnglish (US)
Pages (from-to)1968-1977
Number of pages10
JournalBiophysical Journal
Volume101
Issue number8
DOIs
StatePublished - Oct 19 2011
Externally publishedYes

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Cardiac Myosins
Atomic Force Microscopy
Myosins
Immunoglobulins
Actins
myosin-binding protein C
Sarcomeres
Baculoviridae
Actin Cytoskeleton
Fibronectins
Recombinant Proteins
Proteins

ASJC Scopus subject areas

  • Biophysics

Cite this

Mechanical unfolding of cardiac myosin binding protein-C by atomic force microscopy. / Karsai, Árpád; Kellermayer, Miklós S Z; Harris, Samantha -.

In: Biophysical Journal, Vol. 101, No. 8, 19.10.2011, p. 1968-1977.

Research output: Contribution to journalArticle

Karsai, Árpád ; Kellermayer, Miklós S Z ; Harris, Samantha -. / Mechanical unfolding of cardiac myosin binding protein-C by atomic force microscopy. In: Biophysical Journal. 2011 ; Vol. 101, No. 8. pp. 1968-1977.
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