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Nanomechanics of full-length nebulin: an elastic strain gauge in the skeletal muscle sarcomere.

ABSTRACT: Nebulin, a family of giant modular proteins (MW 700-800 kDa), acts as a F-actin thin filament ruler and calcium-linked regulator of actomyosin interaction. The nanomechanics of full length, native rabbit nebulin was investigated with an atomic force microscope by tethering, bracketing, and stretching full-length molecules via pairs of site-specific antibodies that were attached covalently, one to a protein resistant self-assembled monolayer of oligoethylene glycol and the other to the cantilever. Using this new nanomechanics platform that enables the identification of single molecule events via an unbiased analysis of detachment force and distance of all force curves, we showed that nebulin is elastic and extends to approximately 1 microm by external force up to an antibody detachment force of approximately 300-400 pN. Upon stretching, nebulin unravels and yields force spectra with craggy mountain range profiles with variable numbers and heights of force peaks. The peak spacings, analyzed by the model-independent, empirical Hilbert-Huang transform method, displayed underlying periodicities at approximately 15 and approximately 22 nm that may result from the unfolding of one or more nebulin modules between force peaks. Nebulin may act as an elastic strain gauge that interacts optimally with actin only under appropriate strain and stress. This stretch to match protein ruler may also exert a compressive force that stabilizes thin filaments against stress during contraction. We propose that the elasticity of nebulin is integral and essential in the muscle sarcomere.

SUBMITTER: Yadavalli VK

PROVIDER: S-EPMC2998391 | biostudies-literature | 2009 Jul

REPOSITORIES: biostudies-literature

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Nanomechanics of full-length nebulin: an elastic strain gauge in the skeletal muscle sarcomere.

Yadavalli Vamsi K VK Forbes Jeffrey G JG Wang Kuan K

Langmuir : the ACS journal of surfaces and colloids 20090701 13

Nebulin, a family of giant modular proteins (MW 700-800 kDa), acts as a F-actin thin filament ruler and calcium-linked regulator of actomyosin interaction. The nanomechanics of full length, native rabbit nebulin was investigated with an atomic force microscope by tethering, bracketing, and stretching full-length molecules via pairs of site-specific antibodies that were attached covalently, one to a protein resistant self-assembled monolayer of oligoethylene glycol and the other to the cantilever ...[more]

PMID: 19463013

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Project description:The sarcomere length non-uniformity theory (SLNT) is a widely accepted explanation for residual force enhancement (RFE). RFE is the increase in steady-state isometric force following active muscle stretching. The SLNT predicts that active stretching of a muscle causes sarcomere lengths (SL) to become non-uniform, with some sarcomeres stretched beyond actin-myosin filament overlap (popping), causing RFE. Despite being widely known, this theory has never been directly tested. We performed experiments on isolated rabbit muscle myofibrils (n?=?12) comparing SL non-uniformities for purely isometric reference contractions (I-state) and contractions following active stretch producing RFE (FE-state). Myofibrils were activated isometrically along the descending limb of the force-length relationship (mean?±?1 standard deviation (SD)?=?2.8?±?0.3?µm sarcomere(-1)). Once the I-state was reached, myofibrils were shortened to an SL on the plateau of the force-length relationship (2.4?µm sarcomere(-1)), and then were actively stretched to the reference length (2.9?±?0.3?µm sarcomere(-1)). We observed RFE in all myofibrils (39?±?15%), and saw varying amounts of non-uniformity (1 SD?=?0.9?±?0.5?µm) that was not significantly correlated with the amount of RFE, but through pairwise comparisons was found to be significantly greater than the non-uniformity measured for the I-state (0.7?±?0.4?µm). Three myofibrils exhibited no increase in non-uniformity. Active stretching was accompanied by sarcomere popping in four myofibrils, and seven had popped sarcomeres in the I-state. These results suggest that, while non-uniformities are present with RFE, they are also present in the I-state. Furthermore, non-uniformity is not associated with the magnitude of RFE, and myofibrils that had no increase in non-uniformity with stretch still showed normal RFE. Therefore, it appears that SL non-uniformity is a normal associate of muscle contraction, but does not contribute to RFE following active stretching of isolated skeletal muscle myofibrils.

Dataset Information

Nanomechanics of full-length nebulin: an elastic strain gauge in the skeletal muscle sarcomere.

Publications

Nanomechanics of full-length nebulin: an elastic strain gauge in the skeletal muscle sarcomere.

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