Cell membrane, transport

Based on the lecture by Prof.

Gábor Szabó, 2018

György Vereb, 2020

This lecture:

• Essential Cell Biology: chapters 11-12

All the pages are required, but:

• ion channels, the pumps and mechanisms relevant to

Ca, osmo- and pH regulation, also included in this

chapter of the book, will be discussed in other lectures

• - lecture slides marked by: contain basic

information absolutely required for tests and exams!

!

Lipid

bilayer

5 nm

Functions of the

plasmamembrane:

barrier

transport

signal transduction

Composition:

40-60 % lipid

30-50 % protein

10 % carbohydrate

RevisionMembrane structure and function

Revision

the carbohydrate components of the membrane (from the glycocalyx)

functions:

- surface protection

- cell communication,

recognition

- cell adhesion, extracellular

matrix

Membrane structure: carbohydrates

glycolipids (rare) and glycoproteins (common)

Membrane proteins – function and structure Revision

Inositol

GPI-linked protein

GPI = glycosil-

phosphatydilinositol

Membrane proteins interact with the Cell Cortex(actin meshwork in the cytosol ”below” the membrane)

Red blood cell membrane and its

connections with the cell cortex

Important proteins: actin, ankyrin

(anchor to transmembrane proteins)

spectrin (dimer, forms a mesh)

Intracellular side!!!

!

Molecular complexity of membrane – cell cortex connection

Red blood cell:

Over 50 transmembrane proteinsOutlook

Figure 4. Red cell morphology. Hereditary

spherocytosis (HS; top panel); nonhemolytic

hereditary elliptocytosis (HE; middle panel);

elliptocytes, poikilocytes, and fragmented red cells in

hemolytic HE (bottom panel). BLOOD, 2008, 112(10)

Cell shape is determined by

cell cortex cytoplasmic content

E

S

Hb-S Hb-A

Defects in the vertical spectrin interactions:

spherocytosis (S)

Defects in horizontal interactions:

elliptocytosis (E).

A Info

!

Lipid synthesis and steady-state composition of various membrane

structures

Mitochondria

have bacterial

lipids.

Low cholesterol

in Mitoch., ER,

nuclear envelope

Major

differences

between

organelles

site of synthesis: mostly SER (also Golgi, plasma membrane inner surface)

mammals

yeastsInfo

!

!

!

!

!

1. lipid incorporation into cytoplasmic leaflet

2. flippases/floppases scramblases

3. selective retention

The lipid composition of the cytoplasma

membrane is asymmetric

PS exposure on

the surface of

dying cells: ”eat-

me” signal for the

phagocytes

!

Asymmetric

composition curvature

Lipid shape determines membrane curvature

©2011 by Cold Spring Harbor Laboratory Press

curvature

Info

cylinders (e.g.: PC)

cones (e.g.: PE)

inverted cones

(e.g.: lysoPC)

PC: phosphatidyl choline; PE : phosphatidyl ethanolamine

PS PS

repel

…unless shielded by cholesterol,

so in equilibrium

CHOL/PS/CHOL/PS/CHOL can be flat, but

after extracting cholesterol, PS/PS/PS/PS is curved

Lipid charge also matters…

Hundertwasser,

Vienna

Lipids alone can form complex structures in artifical membranes

Outlook

Cytosolic and exoplasmic surfaces !

Mobility may be restricted by…

(E)(D)

!

Lateral mobility

… clustering / molecular interactions

… and molecular fences

Frye-Edidin expt.

Focal adhesion

Immune synapse

signalosome

hypotonic isotonic

ghostvvt

- Allows diffusion of hydrophobic substances

- Renders membranes deformable

- Explains their regenerative capacity

Deformability – important e.g. in

capillaries

Expreimnt

demonstrating

regenerative

capacity

Right side out

Inside out

Membrane fluidity

3. Energetics– Passive

• Simple diffusion

• Facilitated transport

– Active

• Primary

• secondary

4. Solubility of transported

substance– hydrophylic

– hydrophobic

1. Membrane structure– lipid bilayer

– complete biological membrane

– one or two bilayers

2. Number of different transported

substances and direction of transport– uniport

– Cotransport

– Symport

– Antiport

m

aq

C

CR

Classification of membrane transport from various aspects !

RevisionSimple diffusion through artificial lipid bilayers

RevisionReal (biological) membranes: passive and active

transport

Passive transport

Along electrochemical

gradient

Active transport

Against electrochemical

gradient

Simple

diffusion

Channel transporter/

carrier

Electrochemical

gradient *

* For charged particle, make sure to use ”Electrochemical

gradient”, concentration gradient is wrong!

mechanism

resembles that of

valinomycin, an

ionophore antibiotic

K+

!Facilitated diffusion = Carrier-mediated (passive) transport

E.g. Glucose uniport(see examples in the Biophysics lecture!)

symport antiport

P V F ABCNa/K* Pgp,etc

Vacuolar

Mitochondrial

Secondary active

transporters, carriers

!Main categories of active transport

* The Na/K-ATPase is also

antiport by directionality, but it is

not a coupled/secondary active

transporter; it is a P-type pump.

See next lecture

!ATP- driven pumps. 1. P-type ATPases

P-type ATPases get transiently phosphorylated during their duty cycle

Typical examples are the Na/K-ATPase (see Biophysics lecture also! and the calcium

ATPases (see calcium homeostasis lecture).

!ATP- driven pumps: 2. V and F type ATPases

- V-type ATPases DO NOT get

transiently phosphorylated

- ‚V’ for Vacuole or Vesicle

- Responsible for acidification of

lysosomes

- Against H+ gradient

Pro

ton

gra

dien

tP

roto

n g

rad

ien

t

lysosome Mitoch.

matrix

- F0F1 ATPases DO NOT get

transiently phosphorylated

- Structurally related to V-type

- Generates ATP in mitochondira

- Uses H+ gradient

- Works in reverse as compared to

V-type under physiological

conditions (but can be reversed)

What do you expect to

occur when inhibiting

the Na/K-pump? (with

digitalis or ouabain)

!Intestinal glucose transport

http://academic.brooklyn.cuny.edu/biology/bio4fv/page/sympo.htm

~ 30% of total energy consumption in the cell!

!

Most coupled (secondary active) transport processes depend on the

Na+ gradient, which is built up by the Na/K ATPase

What makes intestinal glucose transport unidirectional (rectified)?

Segregation of specific transport proteins

(„Gram-negative”) bacteria have two cell membranes

outer membrane: porins render it permeable

inner membrane: transporters

Lodish, Fig.

15-15

Alberts, Fig.

11-17

info

Transport of hydrophobic molecules

• Passive transport depends on:

– R (lipid / water partition coeff.)

– i.c. partition - Henderson-Hasselbalch eq.

• Active transport:

– ABC (ATP binding casette) transporters – next lecture

– Other transporters

pH = pK + log(M/M+)

1C

Caq

m

R

Hydrophobic

!

lipid-water partition coeff.

effe

ctiv

eco

nce

ntr

atio

nLipid-water partition coefficient (R) !

For non-charged molecules, lipid-water partition coefficient alone

determines accumulation in the membrane and in the cell. The higher

the R, the more the concentration in the membrane and also in the

cytosol.

General anaesthetics are usually

lipid soluble. Their efficacy is

proportional to their R value, so

the higher their R, the smaller

concentration is sufficient for

reaching an anaesthetic effect.

General anaesthetics can change the conformation of ion channels

through embedding into the membrane and as a result increasing

lateral pressure within the membrane.

hydrostatic

pressure

ether

Ether added to the fishtank

partitions into the cell membrane

(also) of the fish’s neurons, and

inhibits Na+ channels. Incerasing

hydrostatic pressure (e.g. filling

up the tank with a high level of

water) exprimes the ether

molecules from the membrane

and the fish wakes up.

R-NH2

lysosome

pH ≈ 5cytoplasm

pH ≈ 7

R-NH2

R-NH3+

R-NH3+

pH-dependent partitioning !

”Lysosomotropic amines” can accumulate in lysosomes. At physiological pH, these

compounds are mostly unionized and passively diffuse across the lipid bilayers of

organelles. Upon entering the acidic environment of the lysosome they become

predominantly ionized and therefore less able to diffuse out, resulting in their

accumulation. The high cc. (1) is harmful for the lysosomes and (2) sequesters much of

the drug, decreasing its effect on other targets. Local anaesthetics (e.g. lidocaine) and

some cytostatic drugs (e.g. daunorubicine) are examples.

pH = pK + log(M/M+)pK: the pH where the concentration of

neutral and charged forms is equalHenderson-Hasselbalch: determines intracellular

distribution of amphiphilic materials

Daunorubicin accumulation in lysosomes

Blood,Vol 89, No 10 (May 15), 1997: pp 3745-3754

Daunorubicin, an amine that can be protonated, is a DNA-intercalating drug used in

cancer treatment . It exerts its effect partly in the nucleus. Tumor cells with high

lysosomal activity may defer some of its nuclear effect by sequestering it in acidic

lysosomes.

Keywords

• amphiphilic (amphipathic)

compounds

• lipid bilayer

• electrochemical gradient

• lipid-water partitioning

coefficient

• Henderson-Hasselbach equation

and its meaning

• facilitated diffusion

• carrier mediated passive

transport

• passive and active transport

• coupled transport (secondary

active) transport, examples of

(Na/glucose and Na/Ca)

• Na/K pump,

• glucose uniport

• P-type, V-type transporters

!