Related Experiment Video

Updated: Aug 11, 2026

From Constructs to Crystals – Towards Structure Determination of β-barrel Outer Membrane Proteins
09:55

From Constructs to Crystals – Towards Structure Determination of β-barrel Outer Membrane Proteins

Published on: July 4, 2016

Monomer-monomer interactions drive the prepore to pore conversion of a beta-barrel-forming cholesterol-dependent

Eileen M Hotze1, Alejandro P Heuck, Daniel M Czajkowsky

  • 1Department of Microbiology and Immunology, the University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104, USA.

The Journal of Biological Chemistry
|January 19, 2002
PubMed

Insights

A mutant perfringolysin O (PFO) protein gets stuck in a prepore complex. Adding normal PFO triggers the mutant to form a complete pore, showing cooperative binding is key.

Area of Science:

  • Microbiology
  • Biochemistry
  • Molecular Biology

Background:

  • Perfringolysin O (PFO) is a cholesterol-dependent cytolysin.
  • PFO forms large oligomeric pore complexes involving up to 50 PFO molecules.

Purpose of the Study:

  • To investigate the mechanism of PFO pore formation.
  • To identify mutations that trap PFO in a prepore state.
  • To understand the transition from prepore to pore complex.

Main Methods:

  • Site-directed mutagenesis to create PFO(Y181A) mutant.
  • Biochemical assays to analyze PFO oligomerization and pore formation.
  • Characterization of the PFO(Y181A) prepore complex.

Main Results:

  • A PFO mutant (PFO(Y181A)) was identified that forms stable prepore complexes.
  • The PFO(Y181A) prepore complex cannot insert transmembrane beta-hairpins.
  • Incorporation of functional PFO into the PFO(Y181A) prepore induces beta-hairpin insertion and pore formation.
  • The prepore to pore transition is an 'all or none' process, with no partial insertion observed.

Conclusions:

  • Cooperative interactions between PFO monomers drive the transition from prepore to pore.
  • This cooperative mechanism is essential for the formation of the transmembrane beta-barrel structure.
  • The PFO(Y181A) mutant serves as a valuable tool for studying the PFO pore assembly pathway.

More Related Videos

Automated Lipid Bilayer Membrane Formation Using a Polydimethylsiloxane Thin Film08:23

Automated Lipid Bilayer Membrane Formation Using a Polydimethylsiloxane Thin Film

Published on: July 10, 2016

High-throughput Measurement of Plasma Membrane Resealing Efficiency in Mammalian Cells10:07

High-throughput Measurement of Plasma Membrane Resealing Efficiency in Mammalian Cells

Published on: January 7, 2019

Related Concept Videos

Membrane Fluidity01:23

Membrane Fluidity

Cell membranes are composed of phospholipids, proteins, and carbohydrates loosely attached to one another through chemical interactions. Molecules are generally able to move about in the plane of the membrane, giving the membrane its flexible nature called fluidity. Two other features of the membrane contribute to membrane fluidity: the chemical structure of the phospholipids and the presence of cholesterol in the membrane.Fatty acids tails of phospholipids can be either saturated or...
Membrane Fluidity01:26

Membrane Fluidity

Membrane fluidity is explained by the fluid mosaic model of the cell membrane, which describes the plasma membrane structure as a mosaic of components—including phospholipids, cholesterol, proteins, and carbohydrates—that gives the membrane a fluid character.
Mosaic nature of the membrane
The mosaic characteristic of the membrane helps the plasma membrane remain fluid. The integral proteins and lipids exist as separate but loosely-attached molecules in the membrane. The membrane is a relatively...
Porin Insertion in the Outer Mitochondrial Membrane01:12

Porin Insertion in the Outer Mitochondrial Membrane

Porins are beta-barrel proteins translocated to the mitochondrial outer membrane through the TOM complex into the intermembrane space. Porin precursors bind TIM chaperones within the intermembrane space and are guided to the Sorting and Assembly Machinery complex or SAM complex on the outer mitochondrial membrane.
Three models describe the assembly of porins by the SAM complex and their insertion into the outer membrane. Model 1 suggests that porins are assembled outside the SAM channel as the...
Multi-pass Transmembrane Proteins and β-barrels01:09

Multi-pass Transmembrane Proteins and β-barrels

In multi-pass transmembrane proteins, the polypeptide chain crosses the membrane more than once. The transmembrane polypeptide chain either forms an α-helix or β-strand structure. α-Helix containing multi-pass transmembrane proteins are ubiquitous, whereas β-strand containing ones are mainly found in gram-negative bacteria, mitochondria, and chloroplasts.
α-Helix containing multi-pass transmembrane proteins
Multi-pass transmembrane proteins such as G-protein-linked receptors (GPCRs) and...
Mechanisms of Membrane Domain Formation00:59

Mechanisms of Membrane Domain Formation

Different physical properties of lipids and proteins allow them to localize and form distinct islands or domains in the membrane. Some membrane domains are formed due to protein-protein interactions, whereas others are formed due to the presence of specific lipids such as sphingolipids and sterols—for example, large proteins, such as bacteriorhodopsin, aggregate and create distinct domains.
Another mechanism for membrane domain formation involves membrane proteins interacting with cytoskeletal...
Structure of Porins01:21

Structure of Porins

Mitochondria, chloroplasts, and gram-negative bacteria have transmembrane, beta-barrel proteins called porins to mediate the free diffusion of ions and metabolites across the membrane. Mitochondrial porin precursors contain conserved amino acid sequences called beta signals at their C-terminal. Beta signals have a  motif of PoXGXXHyXHy (Po-Polar, X-Any amino acid, G-Glycine, Hy-LargeHydrophobic), which are crucial for precursor recognition to initiate precursor assembly. Beta-barrel precursors...