ABC proteins and the related diseases

Dr. Katalin Goda

Keywords:
active transport, ABC transporters, P-glycoprotein (MDR1, ABCB1),
multidrug resistance, Cystic fibrosis transmembrane conductance
regulator = CFTR=ABCC7, cystic fibrosis, TAP1/TAP2 oligopeptide
transporter, sulfonylurea receptor 1 = SUR1, ABCG2
 REVISION
Active transporters
• Charachteristics:
– Substrate transport against the concentration gradient
or electrochemical potential gradient
– Energy requirement (ATP hydrolysis)
• Types:
– P-type ATPases: Na+/K+-pump, Ca++-pump
– V-type ATPases: H+-ATPase (in the endosomes, lysosome
membrane)
– F-type ATPases: F0F1-ATPase (mitochondria)
– ABC-transporters
 info
ABC transporters are present in species of all kingdoms

48
 !
The basic structure of ABC transporters

• 2 ATP-binding sites (NBDs): binding and

hydrolysis of 2 ATP molecules
• 2 transmembrane domains (TMDs)
collectively forming the substrate binding
sites
• The ATP binding sites are built from evolutionary conserved amino acid sequences
• The ATP molecules are collectively bound by the two ATP binding sites !
C-terminal ATP
binding domain
N-terminal ATP
binding domain
N-terminal ATP C-terminal ATP
binding domain binding domain
ATP

ATP
 ABC transporter categories based on the direction of transport !
Exporters Importers=permeases
• Export of toxic substances (xenobiotics), metabolic • Uptake of amino acids , iron, mono- or
products, antibiotics (antibiotics resistance), drugs, polysacharides, peptides into the bacterial
cytostatics from cells cells
• In both Prokaryotic és Eukaryotic cells • Only in Bacteria
e.g. Periplasmic permeases in gram-negative
• Plasmodium falciparum (Protozoan parasite) is
bacteria
responsible for malaria infection. An ABC
Solute
transporter (Pfmdr) expressed in the plasma
membrane of Plasmodium causes chloroquine Porin
resistance
Outer
membrane
Substrate binding Empty substrate
protein binding protein
Extracellular side

Periplasmic
space

Plasmodium cell

Inner
membrane

chloroquine
ABC importer Cytoplasm
 info
Hydrophobic coating materials (waxes) and anthocyanins are
transported by plant ABC transportres
 Human ABC proteins !
(48 proteins)
• Channel-type ABC proteins:
– Cystic fibrosis transmembrane conductance regulator
CFTR = ABCC7
• Channel regulator:
– SUR1, (CFTR)
• Active transporter:
– Multidrug transporters: Pgp, ABCG2, MRP1
– Phosphatidylcholine flippase: MDR3=ABCB4
– Oligopeptide transporter in the ER: TAP1/TAP2
 Structure of CFTR Cl- -ion channel !
ATP binding and
phosphorylation of the
closed R-domain open
Cl- flow

• Channel-type ABC protein

• ATP binding and hydrolysis dependent
channel gating
• Channel opening is induced by ATP
binding to the NBDs and protein kinase A
ATP hydrolysis
dependent phosphorylation of the R-
domain
• The R domain is an „unstructured”
protein region
 Cystic fibrosis (CF) !
• CFTR is involved in production of sweat, digestive fluids, and mucus
• CF is caused by inactivating mutations in the CFTR Cl--ion channel
expressed in the apical membrane of epithelial cells
• Multiorgan recessive genetic disorder (~ 70 thousands illnesses world-
wide)
• Symptoms affect gastro-intestinal system, lungs and reproductive system
• Serious lung infections, infertility

• Cholera infection: Cholera toxin→…→ adenylil cyclase activation →high c-

AMP level→protein kinase A activation →phosphorylation of the R-
domain of CFTR channels in the intestinal epithelium→Cl- efflux →water
loss (CF carriers have higher chance to survive cholera infection)
!
 info
Effects of different types of mutations on the expression and/or function
of CFTR protein

Wild-type

Abundance
 info
Pharmacological modulation of the defects
caused by mutations of CFTR
 KATP-channel complex !

Hetero-octamer

SUR1: involved in the blood glucose level dependent

regulation of insulin secretion in the beta cells of
pancreas

Increased glucose level in the blood → elevated glucose uptake into the
beta cells→ increased intracellular ATP/ADP ratio →ATP binding to
SUR1/Kir6.2 complex → conformational change → closure of Kir6.2 K+-
channel → depolarization → elevated intracellular Ca++ level → insulin
secretion
 !
Regulation of insulin secretion by the KATP-channel complex

Low blood glucose level: High blood glucose level, (sulfonylurea analogs):
NO insulin
secretion

KATP-channel KATP-channel
OPEN CLOSED

Loss of function mutation →HYPOGLICAEMIA Gain of function mutation →DIABETES

NO insulin
secretion

KATP-channel
KATP-channel
CLOSED
OPEN
 P-glycoprotein (Pgp, MDR1, ABCB1) !
substrates
Expression:, small intestine,, blood-brain
barrier, placenta, blood-testis barrier,
tumor cells, stem cells
Plasma Substrates: toxic compounds, amphiphylic
membrane and lipophylic xenobiotics, chemo-
therapeutic drugs
cytoplasm Knock-out: Blood-brain barrier insufficiency
Mechanism: membrane → water phase
Significance: tumor and AIDS
chemotherapy
ATP

Blood-brain barrier Placenta

Glia
Maternal side

Basolateral membrane
 Multidrug resistance !
Pgp+ tumor Pgp+ tumor Pgp- tumor
+ inhibitor NO Inhibitor

• A daganatsejtek plazmamembránjában kifejeződő széles szubsztrát Pgp+

tumor
• Inhibitor nélkül
• spektrumú ABC transzporter fehérjék okozzák
• P-glikoprotein (Pgp), ABCG2 és multidrog rezisztencia protein 1(MRP1)
•• Resistance
Őssejtekenagainst chemically and
is kifejeződhetnek ezekfunctionally unrelated drugs
a transzporterek
• Cytotoxic drugs do not reach the cell killing dose in tumor cells
• It is caused by the expression of ABC transporters (with wide substrate
spectrum) in the plasma membrane of cancer cells: P-glycoprotein
(Pgp=MDR1=ABCB1), ABCG2 and multidrug resistance protein
1(MRP1=ABCC1)
• Their expression is induced by the chemotherapy or they are
intrinsically expressed

Tumor cells,
 What is common? info

Leonardo da Vinci Henry VIII Isaac Newton Franklin Benjamin

Ludwig van Beethowen Charles Darwin Luciano Pavarotti

 ABCG2 and gout !
Homodimer

• ABC transporter with wide substrate spectrum

• Expressed in tissue barriers and stem cells
• Half transporter→forms homodimers
• Endogenous substrate: uric acid

podagra

• Mutations decreasing the function of ABCG2 (e.g. Q141K) → elevated urate level
in the blood → increased risk of gout
• Symptoms of gout: Uric acid precipitates and forms deposits (tophi) in joints and
in the surrounding tissues →Recurrent attack of accute inflammatory arthritis
 ABC-pumps in the liver

cholesterol
ABCG5/G8

ABCG2

• Major components of bile: phosphatidylcholine, cholesterol and bile acids are

transported by ABC transporters
• Detoxification in the liver = conjugation of toxic compounds with glutathion and !
glucuronide transport to the bile by ABC transporters
 TAP1/TAP2 is an olygopeptide transporter in the ER membrane !

TAP1/TAP2

Cellular and certain viral proteins degraded in the proteasomes to oligopeptides are
pumped into the lumen of ER by TAP1/TAP2 transporter. In the ER lumen they bind
to MHCI molecules and the complex is transported to the plasma membrane to be
exhibited to cytotoxic T-cells.
 Diseases caused by mutations in ABC
transporters
• CFTR (ABCC7), chloride-ion channel – cystic fibrosis
• BCRP (ABCG2), urate transporter – gout
• TAP1/TAP2, oligopeptide transporter – immundeficiency
• SUR1 (ABCC8), potassium channel regulator
– gain of function mutation – diabetes
– loss of function mutation – hyperinsulinaemic hypoglicaemia
• ABCA1, cholesterol and phospholipid transporter involved
in the formation of HDL particles – Tangier disease
• ABCA4, transport of all-trans retinaldehyde derivatives in
rods and cones – age related macula degeneration,
Stargardt disease
Materials for Seminar
Describe the intestinal glucose uptake. Which transporters
take part in the process?
How the insulin secretion is regulated in the beta cells?
Describe how the blood glucose level is regulated.
• Na/glucose symporter (secondary active transport) in the apical part of
intestinal epithelial cells – glucose uptake from the gut lumen
• Glut1 (glucose uniport) in the basolateral surface of intestinal epithelial
cells (passive transport) – glucose transport from the epithelial cells to the
blood
• Regulation of the insulin secretion: Increased glucose level in the blood →
elevated glucose uptake into the beta cells→ increased intracellular
ATP/ADP ratio →ATP binding to SUR1 → conformational change → closure
of Kir6.2 K+-channel → depolarization → elevated intracellular Ca++ level →
insulin secretion (typical example for regulated secretion)
• Insulin binding to the insulin receptor (expressed on the surface of muscle
cells)→……→Glut4 expressing vesicles fuse with the cell membrane →
elevated Glut4 expression →glucose uptake to the muscle cells →decrease
of blood glucose level (details see in Signaling II lecture)
 Describe the studied ABC proteins answering the following
questions.

P-glycoprotein (ABCB1), ABCG2, CFTR (ABCC7), SUR1, TAP1/TAP2

• In which tissues are they expressed?

• Describe their subcellular expression site.
• Do they belong to the channel, channel regulator or
active transporter categories of human ABC
proteins?
• Which biological processes they play a role?
• Name and describe the pathological processes
related to the above ABC proteins.
Describe the function, tissue expression and physiological role of CFTR
protein.
Which disease is related to the mutations of CFTR protein? What are the
symptoms of this disease? How this disease can be treated?

• Chloride channel in the apical membrane of epithelial

cells
• Sites of expression: airways, pancreas, intestinal
epithelium, sweat glands, reproductive organs
• Function: mucus production
• Cystic fibrosis (CF): recurrent airway infections,
pneumonia, digestion problems, infertility
• Treatment: in case of F508 deletion „corrector”
molecules support the normal folding of the protein