Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Experiment Videos

Clathrin assembly proteins: affinity purification and a model for coat assembly.

J H Keen1

  • 1Fels Research Institute, Temple University School of Medicine, Philadelphia, Pennsylvania 19140.

The Journal of Cell Biology
|November 1, 1987
PubMed
Summary

Researchers identified two distinct assembly proteins (APs), AP-1 and AP-2, from bovine brain coated vesicles. These proteins, AP-1 and AP-2, are crucial for clathrin coat assembly and influence the size of the resulting structures.

Related Concept Videos

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

The actin-binding protein Hip1R associates with clathrin during early stages of endocytosis and promotes clathrin assembly in vitro.

The Journal of cell biology·2001
Same author

The class II phosphoinositide 3-kinase C2alpha is activated by clathrin and regulates clathrin-mediated membrane trafficking.

Molecular cell·2001
Same author

Selective recruitment of arrestin-3 to clathrin coated pits upon stimulation of G protein-coupled receptors.

Journal of cell science·2000
Same author

The class II phosphoinositide 3-kinase PI3K-C2alpha is concentrated in the trans-Golgi network and present in clathrin-coated vesicles.

The Journal of biological chemistry·2000
Same author

Spatial control of coated-pit dynamics in living cells.

Nature cell biology·1999
Same author

Phosphoinositide-AP-2 interactions required for targeting to plasma membrane clathrin-coated pits.

The Journal of cell biology·1999

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Biochemistry

Background:

  • Coated vesicles are essential for intracellular transport.
  • Assembly proteins (APs) play a role in the formation of clathrin coats.
  • Previous studies have described assembly proteins involved in coated vesicle formation.

Purpose of the Study:

  • To fractionate and characterize assembly proteins from bovine brain coated vesicles.
  • To investigate the role of different AP fractions in clathrin coat assembly.
  • To elucidate the mechanism of AP-mediated clathrin coat formation.

Main Methods:

  • Clathrin-Sepharose affinity chromatography was used to fractionate APs.
  • Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was employed to determine the molecular weights of protein components.

Related Experiment Videos

  • Low angle laser light scattering was used to determine the molecular weight of native AP-2.
  • In vitro assembly assays were performed with clathrin and AP fractions.

    • Main Results:

    • Two distinct AP fractions, AP-1 and AP-2, with different subunit compositions and molecular weights were isolated.
    • Both AP-1 and AP-2 induced clathrin coat formation in vitro.
    • AP-1 formed slightly smaller coats (72 nm) compared to AP-2 (79 nm).
    • AP-2 was determined to be a functional dimer, suggesting bivalent clathrin binding.
    • A model for AP-mediated coat assembly was proposed, involving AP-2 bridging clathrin trimers.

    Conclusions:

    • A family of assembly proteins exists within cells, with at least AP-1 and AP-2 involved in clathrin coat assembly.
    • AP-2's bivalent nature is proposed to be key in initiating and stabilizing clathrin lattice formation.
    • The interaction of APs with clathrin trimers influences coat structure size and curvature, potentially contributing to vesicle formation during endocytosis.