unit 4- membrane proteins

Percentage of cellular proteins that are membrane-associated

$$\sim 30\%$$

Percentage of current therapeutic targets that are integral membrane proteins

$$\sim 60\%$$

Percentage of membrane weight accounted for by protein

$$\sim 50\%$$

Approximate ratio of lipids per membrane protein

$\sim 50\text{--}100$ (proteins are significantly larger than lipids)

Fundamental structural elements of biological membranes

Lipids

Membrane components that carry out specific dynamic functions

Proteins

Structural basis for membrane "sidedness"

Proteins attach to and insert into the lipid bilayer asymmetrically.

Three major classes of membrane proteins

Integral, lipid-anchored, and peripheral proteins.

Integral membrane proteins

Proteins that penetrate and pass through the lipid bilayer as single-pass or multipass structures.

Amphipathic organization of integral membrane proteins

Hydrophobic domains anchor them within the interior of the lipid bilayer, while hydrophilic regions form functional domains exposed to the aqueous exterior.

Interactions that seal the membrane barrier around integral proteins

van der Waals interactions with neighboring lipids, which preserve the permeability barrier.

Monotopic membrane proteins

Integral proteins that embed into one leaflet of the membrane without emerging on the opposite side.

Membrane orientation resulting from an odd number of transmembrane domains

"In-out" orientation, where the protein termini reside on opposite sides of the bilayer.

Membrane orientation resulting from an even number of transmembrane domains

"In-in" or "out-out" orientation, where both protein termini reside on the same side of the bilayer.

Transmembrane proteins

Proteins that span across the lipid bilayer with functional portions exposed on both the extracellular and cytosolic faces.

Location of hydrophobic regions in transmembrane proteins

In the interior of the lipid bilayer, in direct contact with nonpolar fatty acid tails.

Location of hydrophilic regions in transmembrane proteins

Exposed to the aqueous environment on either side of the membrane.

Chemical properties of hydrophobic amino acids

Uncharged and nonpolar.

Chemical properties of hydrophilic amino acids

Charged and polar.

Attachment mechanism of lipid-anchored proteins

The polypeptide is covalently linked to a lipid embedded within the bilayer, while the protein itself remains outside the membrane core.

Three primary types of lipid-anchored proteins

GPI-anchored, fatty acid-anchored, and isoprenylated proteins.

Full chemical name of GPI

Glycosylphosphatidylinositol

Subcellular localization of GPI-anchored proteins

External (extracellular) leaflet of the plasma membrane.

Fatty acids predominantly utilized for fatty acid anchoring

Myristic acid and palmitic acid.

Isoprenylation

The covalent attachment of multiple $5$-carbon isoprene units to a protein.

Lipid anchor formed by three isoprene units ($15$ carbons)

Farnesyl

Lipid anchor formed by four isoprene units ($20$ carbons)

Geranylgeranyl

Class of signaling molecules where isoprenylation is common

Small GTPases

Attachment mechanism of peripheral membrane proteins

Weak electrostatic and non-covalent interactions.

Subcellular location of peripheral membrane proteins

Entirely outside the lipid bilayer, localized to either the extracellular or cytoplasmic surface.

Solubilization characteristics of peripheral membrane proteins

Easily solubilized without disrupting the structural integrity of the lipid bilayer.

Binding partners of peripheral membrane proteins

Integral membrane proteins or charged/polar lipid head groups.

Dynamic nature of peripheral membrane protein association

Transient and reversible, modulated by changing cellular conditions and signaling events.

Biochemical conditions used to extract peripheral membrane proteins

High ionic strength (high salt) or extreme pH, which disrupt non-covalent electrostatic interactions.

Transport function of membrane proteins

Facilitating the movement of ions (e.g., establishing $H^+$ gradients across mitochondria, lysosomes, and chloroplasts) and large polar molecules (sugars, amino acids, nucleotides).

Linker function of membrane proteins

Connecting structural proteins on the cytosolic face to macromolecules on the noncytosolic face.

Integrins (as linker proteins)

Proteins that link the intracellular actin cytoskeleton to extracellular matrix components.

Receptor function of membrane proteins

Detecting specific chemical signals on one face of the membrane and transducing that signal across to the opposing face.

Epidermal growth factor receptor (EGFR)

A transmembrane receptor that binds extracellular EGF to trigger intracellular signaling; frequently mutated in various human cancers.

Enzymatic function of membrane proteins

Catalyzing specific biochemical reactions directly at the cytosolic or extracellular surface of the membrane.

Adenylyl cyclase (as a membrane-associated enzyme)

A membrane-bound enzyme that catalyzes the synthesis of cyclic AMP from ATP.

Predominant secondary structure utilized by proteins to span the lipid bilayer

$\alpha$-helix

Reason carbonyl oxygen carries a partial negative charge in peptide bonds

Oxygen is more electronegative than carbon and pulls shared electrons more strongly.

Reason amide hydrogen carries a partial positive charge in peptide bonds

Nitrogen is more electronegative than hydrogen and draws shared electron density toward itself.

Hydrogen-bonding scheme within an $\alpha$-helix

The carbonyl oxygen of residue $i$ forms a hydrogen bond with the amide hydrogen of residue $i+4$.

Residues per complete turn in an $\alpha$-helix

$3.6$ residues per turn

Rise (translation) per complete turn in an $\alpha$-helix

$$0.54\,\text{nm}$$

Amino acid excluded from $\alpha$-helices due to its rigid ring structure

Proline (its cyclic side chain disrupts the spiral conformation).

Side chain characteristics of a single-pass transmembrane $\alpha$-helix

Exclusively hydrophobic and nonpolar, allowing favorable packing with surrounding lipid tails.

Primary physiological role of single hydrophobic transmembrane helices

Receptors for extracellular signaling ligands (transducing binding events to intracellular domains).

Amphipathic $\alpha$-helix

An $\alpha$-helix possessing one hydrophobic face and one charged or polar hydrophilic face.

Membrane protein architecture where amphipathic helices are predominantly found

Multipass transmembrane proteins.

Structure assembled by clustered amphipathic helices within a membrane

A hydrophilic central pore lined by polar residues, surrounded by hydrophobic faces contacting fatty acid tails.

Molecules transported through amphipathic helical pores

Large polar molecules (amino acids, nucleotides, sugars) and small charged ions.

Membrane-spanning secondary structure alternative to the $\alpha$-helix

$\beta$-sheet organized into a $\beta$-barrel.

Difference in hydrogen bonding between $\beta$-sheets and $\alpha$-helices

$\beta$-sheet hydrogen bonds form between residues that can be distant in the primary sequence, rather than strictly between residues $i$ and $i+4$.

Relative strand orientations in a $\beta$-sheet

Parallel or antiparallel.

Periodicity (repeat distance) of an extended $\beta$-strand

$0.7\,\text{nm}$ (due to its pleated, kinked conformation).

Primary stabilizing force between strands in a $\beta$-sheet

Hydrogen bonds between backbone peptide groups of neighboring strands.

$\beta$-barrel

A cylindrical protein motif formed by rolling one or more $\beta$-sheets into a closed barrel.

Side chain orientation on the exterior of a transmembrane $\beta$-barrel

Hydrophobic side chains directed outward to interact with membrane lipid tails.

Side chain orientation lining the interior lumen of a transmembrane $\beta$-barrel

Hydrophilic side chains directed inward to form a water-filled pore.

Physiological role of transmembrane $\beta$-barrel channels

Selective transport of large polar molecules and small charged ions.

Requirement for solubilizing transmembrane integral proteins

Chemical disruption of the lipid bilayer's hydrophobic interactions.

Reagents most commonly used to solubilize integral membrane proteins

Detergents

Detergent

A small amphipathic molecule containing a single hydrophobic tail and a polar or charged hydrophilic head.

Two most widely used detergents in membrane biochemistry

Sodium dodecyl sulfate (SDS) and Triton X-100.

Full chemical name of SDS

Sodium dodecyl sulfate

Chemical classification and strength of SDS

Strong ionic (denaturing) detergent containing an ionized sulfate head group.

Chemical classification and strength of Triton X-100

Mild non-ionic (non-denaturing) detergent containing an uncharged polar head group.

Structural difference between detergent and membrane lipid tails

Detergents possess a single hydrophobic tail, whereas membrane lipids possess two hydrophobic tails.

Geometric shape of a detergent monomer

Cone

Geometric shape of a typical membrane lipid

Cylinder

Self-assembly structure formed spontaneously by cylindrical membrane lipids in water

Lipid bilayers

Self-assembly structure formed spontaneously by conical detergent molecules in water

Micelles

Molecular outcome when detergent is present in great excess relative to membrane proteins

Hydrophobic detergent tails bind the protein's transmembrane segments and lipid tails, displacing lipids and solubilizing the protein.

Mechanism by which detergent-protein complexes remain dissolved in aqueous solution

Water molecules structure favorably around the hydrophilic detergent head groups that envelop the hydrophobic domains of the protein.

Lateral mobility of membrane proteins

Proteins can diffuse laterally within the lipid bilayer, though their movement is often constrained into discrete membrane domains.

Mechanism that restricts the lateral diffusion of membrane proteins

Specific protein-protein interactions and tethering complexes.

Four mechanisms that restrict membrane protein lateral mobility

(A) Self-aggregation into large complexes; (B) tethering to extracellular matrix components; (C) anchoring to the intracellular cytoskeleton; (D) binding to membrane proteins on an adjacent cell.

Extracellular matrix component that restricts membrane protein mobility

Collagen

Example of membrane protein restriction via self-aggregation

Crystalline patches of bacteriorhodopsin in the purple membrane of Halobacterium halobium.

Example of membrane protein restriction via cytoskeletal anchoring

Erythrocyte (red blood cell) membrane proteins anchored to the spectrin cytoskeleton.

Cell Adhesion Molecules (CAMs)

Transmembrane proteins that mediate physical binding between cells or between cells and the extracellular matrix.

Three major classes of CAMs

Integrins, selectins, and cadherins.

Homophilic CAM interaction

Binding between identical adhesion molecules located on opposing cell surfaces.

Heterophilic CAM interaction

Binding between different adhesion molecules or ligands on opposing cell surfaces.

Cause of variable lateral diffusion coefficients ($D$) among membrane proteins

Differences in protein immobilization resulting from tethering or molecular crowding.

Physical membrane property governing the rate of lateral protein diffusion

Membrane fluidity

Lipid raft

A specialized, dynamic membrane microdomain enriched in specific lipids and signaling proteins.

Lipid components enriched within plasma membrane lipid rafts

Cholesterol and glycosphingolipids containing long, saturated hydrocarbon tails.

Physical properties of lipid rafts relative to the surrounding membrane

More rigid (liquid-ordered state) and physically thicker.

Protein class preferentially enriched within lipid rafts

GPI-anchored proteins

Membrane protein class anchored solely through non-covalent electrostatic interactions

Peripheral membrane proteins

Membrane protein class anchored through a covalently linked lipid moiety

Lipid-anchored proteins

Membrane protein class that directly spans the hydrophobic interior of the bilayer

Integral membrane proteins

Membrane protein class strictly requiring detergent disruption for biochemical extraction

Integral membrane proteins

Membrane protein class extractable using high ionic strength or extreme pH

Peripheral membrane proteins

Transmembrane helix motif capable of assembling into a functional aqueous pore

Amphipathic $\alpha$-helix (in multipass proteins)

Transmembrane motif whose outward-facing side chains are entirely hydrophobic

Single-pass transmembrane $\alpha$-helix