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September 1, 1991Proceedings of the National Academy of SciencesOpen Access

Site-directed mutations of Dictyostelium actin: disruption of a negative charge cluster at the N terminus.

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Population

Dictyostelium cells expressing mutant actins

Comparison

Site-directed mutagenesis replacing aspartic… vs Wild-type actin

Design

Preclinical

Authors

KSKeita SutohNihon Medi-Physics (Japan)MAMitsushige AndoShiga Medical CenterKSKeita SutohNihon Medi-Physics (Japan)

Discussion

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Implication

These kinetic defects in actin-myosin interaction leave open any role in human cardiomyopathy; further translational studies needed before clinical relevance.

Key Points

  • To investigate how site-directed mutations of the N-terminal negative charge cluster of Dictyostelium actin affect its interaction with myosin.
  • Replaced aspartic acid residues with histidine residues in the actin gene via site-directed mutagenesis.
  • Expressed mutant actins in Dictyostelium cells and purified them using HPLC.
  • Conducted in vitro assays to compare functional properties of mutant and wild-type actins.
  • Mutant actins exhibited slowed sliding movement in actin filaments driven by myosin.
  • The maximum turnover rate of myosin's ATPase activity significantly dropped, while the apparent affinity for actin remained unchanged.

Structured PICO

P
Population
Dictyostelium cells expressing mutant actins
I
Intervention
Site-directed mutagenesis replacing aspartic acid residues with histidine residues in the N-terminal negative charge cluster of Dictyostelium actin
C
Comparator
Wild-type actin
O
Outcome
In vitro sliding movement of actin filaments driven by myosin and actin-activated ATPase activity of myosinsurrogate

The N-terminal negative charge cluster of actin is essential for the ATP-dependent actin-myosin interaction.

Cite This Study

Sutoh et al. (1991) studied this question.

synapsesocial.com/papers/6a13ca003f9a9dbf1d39e3behttps://doi.org/10.1073/pnas.88.17.7711
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Also Consider

Synapse has enriched 5 closely related papers on similar clinical questions. Consider them for comparative context:

  1. 1Complete Amino-Acid Sequence of Actin of Rabbit Skeletal Muscle1973 · 405 citations
  2. 2The Initial Phosphate Burst in ATP Hydrolysis by Myosin and Subfragment-1 as Studied by a Modified Malachite Green Method for Determination of Inorganic Phosphate1986 · 299 citations
  3. 3Expression and Characterization of a Functional Myosin Head Fragment in Dictyostelium discoideum1989 · 76 citations
  4. 4Expression of actin in Escherichia coli. Aggregation, solubilization, and functional analysis.1990 · 43 citations
  5. 5Identification of myosin-binding sites on the actin sequence1982 · 327 citations