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Related Experiment Videos

Stabilization of flagellar filaments by HAP2 capping.

Zoltán Diószeghy1, Péter Závodszky, Keiichi Namba

  • 1Institute of Enzymology, Hungarian Academy of Sciences, Karolina u. 29, H-1113 Budapest, Hungary.

FEBS Letters
|June 16, 2004
PubMed
Summary

Bacterial flagellar filaments are stabilized by a hook associated protein 2 (HAP2) cap. This HAP2 cap enhances filament stability against heat, aiding in structural analysis.

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Area of Science:

  • Microbiology
  • Structural Biology
  • Biochemistry

Background:

  • Bacterial flagellar filaments are essential for motility.
  • Self-assembly of flagellin forms these filaments.
  • A capping structure, hook associated protein 2 (HAP2), is involved in filament growth.

Purpose of the Study:

  • To investigate the role of HAP2 in flagellar filament stability.
  • To compare the stability of native and reconstituted flagellar filaments with and without HAP2.
  • To develop a method for isolating native, HAP2-capped filaments for structural studies.

Main Methods:

  • Isolation of native bacterial flagellar filaments.
  • Thermal denaturation assays to measure melting temperature.
  • Proteolysis assays to assess stability.

Related Experiment Videos

  • In vitro reconstitution of flagellar filaments with HAP2.

    • Main Results:

    • Native filaments with intact HAP2 caps showed increased thermal stability (ΔTm = 4°C) compared to non-capped filaments.
    • HAP2 binding stabilized reconstituted filaments, but these complexes were less stable than native assemblies.
    • Reconstituted filament-HAP2 complexes were sensitive to proteolysis and depolymerized rapidly at elevated temperatures.
    • A procedure was developed to isolate perfectly capped native filaments.

    Conclusions:

    • The HAP2 cap significantly enhances the thermal stability of bacterial flagellar filaments.
    • In vitro reconstitution methods often fail to produce native-like stable filament-HAP2 complexes.
    • The developed isolation procedure enables high-resolution structural analysis of native, HAP2-capped flagellar filaments.