Cross-Species Mechanical Fingerprinting of Cardiac Myosin Binding Protein-C

• Published in: Biophysical journal Vol. 104; no. 11; pp. 2465 - 2475
• Main Authors: Karsai, Árpád, Kellermayer, Miklós S.Z., Harris, Samantha P.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 04.06.2013; Biophysical Society; The Biophysical Society
• Subjects: Animals; atomic force microscopy; Binding sites; Carrier Proteins - chemistry; Carrier Proteins - metabolism; Cattle; Cells; fibronectins; Gene expression; heart; Humans; immunoglobulins; insects; mechanical loads; Mechanical Phenomena; mechanical properties; Mice; Models, Molecular; Molecular Machines, Motors, and Nanoscale Biophysics; Monte Carlo Method; myosin; Protein folding; Protein Structure, Tertiary; Protein Unfolding; Proteins; Recombinant Proteins - chemistry; Recombinant Proteins - metabolism; Species Specificity
• ISSN: 0006-3495; 1542-0086
• DOI: 10.1016/j.bpj.2013.04.027

Cardiac myosin binding protein-C (cMyBP-C) is a member of the immunoglobulin (Ig) superfamily of proteins and consists of 8 Ig- and 3 fibronectin III (FNIII)-like domains along with a unique regulatory sequence referred to as the MyBP-C motif or M-domain. We previously used atomic force microscopy to investigate the mechanical properties of murine cMyBP-C expressed using a baculovirus/insect cell expression system. Here, we investigate whether the mechanical properties of cMyBP-C are conserved across species by using atomic force microscopy to manipulate recombinant human cMyBP-C and native cMyBP-C purified from bovine heart. Force versus extension data obtained in velocity-clamp experiments showed that the mechanical response of the human recombinant protein was remarkably similar to that of the bovine native cMyBP-C. Ig/Fn-like domain unfolding events occurred in a hierarchical fashion across a threefold range of forces starting at relatively low forces of ∼50 pN and ending with the unfolding of the highest stability domains at ∼180 pN. Force-extension traces were also frequently marked by the appearance of anomalous force drops suggestive of additional mechanical complexity such as structural coupling among domains. Both recombinant and native cMyBP-C exhibited a prominent segment ∼100 nm-long that could be stretched by forces <50 pN before the unfolding of Ig- and FN-like domains. Combined with our previous observations of mouse cMyBP-C, these results establish that although the response of cMyBP-C to mechanical load displays a complex pattern, it is highly conserved across species.