Molecular basis of tropomyosin binding to tropomodulin, an actin-capping protein

Alla S Kostyukova; Sarah E Hitchcock-Degregori; Norma J Greenfield

doi:10.1016/j.jmb.2007.05.084

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Molecular basis of tropomyosin binding to tropomodulin, an actin-capping protein

Journal article

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Peer reviewed

Molecular basis of tropomyosin binding to tropomodulin, an actin-capping protein

Alla S Kostyukova, Sarah E Hitchcock-Degregori and Norma J Greenfield

Journal of molecular biology, Vol.372(3), pp.608-618

09/21/2007

DOI: https://doi.org/10.1016/j.jmb.2007.05.084

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https://hdl.handle.net/2376/116006

PMCID: PMC2134803

PMID: 17706248

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https://doi.org/10.1016/j.jmb.2007.05.084View

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Abstract

Protons

Amino Acid Sequence

Actin Cytoskeleton - metabolism

Magnetic Resonance Spectroscopy

Gelsolin - metabolism

Cross-Linking Reagents - pharmacology

Molecular Sequence Data

Mutant Proteins - metabolism

Nitrogen Isotopes

Amino Acids

Tropomodulin - metabolism

Tropomodulin - chemistry

Peptide Fragments - chemistry

Sequence Alignment

Animals

Protein Isoforms - metabolism

Protein Isoforms - chemistry

Chickens

Protein Binding - drug effects

Tropomyosin - chemistry

Circular Dichroism

Tropomyosin - metabolism

The tropomodulin (Tmod) family of proteins that cap the pointed, slow-growing end of actin filaments require tropomyosin (TM) for optimal function. Earlier studies identified two regions in Tmod1 that bind the N terminus of TM, though the ability of different isoforms to bind the two sites is controversial. We used model peptides to determine the affinity and define the specificity of the highly conserved N termini of three short, non-muscle TMs (alpha, gamma, delta-TM) for the two Tmod1 binding sites using circular dichroism spectroscopy, native gel electrophoresis, and chemical crosslinking. All TM peptides have high affinity for the second Tmod1 binding site (within residues 109-144; alpha-TM, 2.5 nM; gamma-TM, delta-TM, 40-90 nM), but differ >100-fold for the first site (residues 1-38; alpha-TM, 90 nM; undetectable at 10 microM, gamma-TM, delta-TM). Residue 14 (R in alpha; Q in gamma and delta) and, to a lesser extent, residue 4 (S in alpha; T in gamma and delta) are primarily responsible for the differences. The functional consequence of the sequence differences is reflected in more effective inhibition of actin filament elongation by full-length alpha-TMs than gamma-TM in the presence of Tmod1. The binding sites of the two Tmod1 peptides on a model TM peptide differ, as defined by comparing (15)N,(1)H HSQC spectra of a (15)N-labeled model TM peptide in both the absence and presence of Tmod1 peptide. The NMR and CD studies show that there is an increase in alpha-helix upon Tmod1-TM complex formation, indicating that intrinsically disordered regions of the two proteins become ordered upon binding. A model proposed for the binding of Tmod to actin and TM at the pointed end of the filament shows how the Tmod-TM accentuates the asymmetry of the pointed end and suggests how subtle differences among TM isoforms may modulate actin filament dynamics.

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Title: Molecular basis of tropomyosin binding to tropomodulin, an actin-capping protein
Creators: Alla S Kostyukova - Department of Neuroscience and Cell Biology, Robert Wood Johnson Medical School, 675 Hoes Lane, Piscataway, NJ 08854, USA
Sarah E Hitchcock-Degregori
Norma J Greenfield
Publication Details: Journal of molecular biology, Vol.372(3), pp.608-618
Academic Unit: Chemical Engineering and Bioengineering, School of
Publisher: England
Grant note: GM63257 / NIGMS NIH HHS GM36326 / NIGMS NIH HHS R01 GM036326-19 / NIGMS NIH HHS R01 GM063257 / NIGMS NIH HHS R01 GM036326 / NIGMS NIH HHS R01 GM063257-17 / NIGMS NIH HHS
Identifiers: 99900547987301842
Language: English
Resource Type: Journal article