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March 1, 1988The Journal of Cell BiologyOpen Access

Probing actin polymerization by intermolecular cross-linking.

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Population

Globular monomeric actin (G-actin) in vitro

Comparison

N,N'-1,4-phenylenebismaleimide vs Unmodified G-actin

Design

Preclinical

Authors

RMRobert C. MillonigUniversity of BaselHSHope SalvoScience Applications International Corporation (United States)UAU. AebiJohns Hopkins University

Discussion

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Implication

No immediate clinical implications; leaves open the in vivo relevance of LD-to-UD transition in actin dynamics.

Key Points

  • To investigate the early oligomeric intermediates and dimerization events during the assembly of monomeric G-actin into filamentous actin.
  • Treated monomeric G-actin with the bifunctional sulfhydryl cross-linking reagent N,N'-1,4-phenylenebismaleimide across varying ionic conditions.
  • Assayed polymerization kinetics and filament formation using SDS-PAGE, N-(1-pyrenyl)iodoacetamide fluorescence enhancement, and electron microscopy.
  • Isolated specific cross-linked dimer species (lower dimer [LD] and upper dimer [UD]) to evaluate their competency for filament assembly and their impact on nucleation kinetics.
  • Early polymerization induces significant formation of an initial 86-kD lower dimer (LD), which later diminishes as an assembly-competent 115-kD upper dimer (UD) appears alongside filament growth.
  • Purified LD cannot assemble into filaments or alter monomer polymerization kinetics, matching symmetries found in crystalline sheets rather than helical filaments.
  • Purified UD directly forms normal actin filaments and accelerates assembly by shortening the kinetic lag phase in a concentration-dependent manner.

Structured PICO

P
Population
Globular monomeric actin (G-actin) in vitro
I
Intervention
N,N'-1,4-phenylenebismaleimide (bifunctional sulfhydryl cross-linking reagent)
C
Comparator
Unmodified G-actin
O
Outcome
Oligomeric state of actin and filament formationsurrogate

Identifies distinct dimer intermediates (LD and UD) during actin polymerization, demonstrating that the UD acts to shorten the lag phase of filament assembly.

Cite This Study

Millonig et al. (1988) studied this question.

synapsesocial.com/papers/6a088e3f7f7fcd1344ddec8bhttps://doi.org/10.1083/jcb.106.3.785
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Also Consider

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

  1. 1Isolation and characterization of covalently cross-linked actin dimer.1981 · 51 citations
  2. 2F-actin is intermolecularly crosslinked by N,N'-p-phenylenedimaleimide through lysine-191 and cysteine-374.1984 · 83 citations
  3. 3Mechanical properties of actin.1985 · 98 citations
  4. 4The primary structure of actin from rabbit skeletal muscle. Five cyanogen bromide peptides, including the NH2 and COOH termini.1975 · 72 citations
  5. 5Characterization of sulfhydryl groups of actin1969 · 92 citations