Lecture 4

Question 1

P-type ATPases

Answer 1

Consist of large alpha (200kDa) and associated beta subunits

Pump cations

Inhibited by orthovanadate

ATP donates gamma-phosphate to aspartate during cation pumping

Question 2

Na+/K+ ATPase

Answer 2

Located in plasma membrane of animal cells

3Na+/2K+ per ATP hydrolysed

Maintains high K+/low Na+ in cytosol
Maintains Na+ electrochemical gradient

Question 3

Structure of Na+/K+ ATPase

Answer 3

2 alpha (112kDa) and 2 beta (34kDa) subunits

Question 4

Fungal and Plant H+ ATPase

Answer 4

Location - Plasma membrane of plants and fungi

Stochimetry of 1H+ per ATP

Expels excess H+

H+ electrochemical gradient maintained for H+ coupled transport

Mainatains negative transmembrane voltage

Regulates cytosolic pH

Loosens cell wall by acidification

Question 5

Structure of fungal and plant H+ ATPase

Answer 5

1 alpha subunit (112kDa)

Question 6

Sarcoplasmic endoreticulum Ca2+ ATPase

Answer 6

• Located in sarcoplasmic reticulum in muscle cell cytoplasm
• 2Ca2+/ATP

Restores low Ca2+ after muscles contract

1 alpha subunit (112kDa) - 3 isoforms

Question 7

Name inhibitor of sarcoplasmic endoreticulum Ca2+ ATPase

Answer 7

Thapsigargin

Question 8

Plasma membrane Ca2+ ATPase

Answer 8

Location is plasma membrane in animals, plants, fungi

1-2Ca2+/ATP (in exchange for H+)

Maintains low cytosolic calcium ion as otherwise cytotoxic
Cell signalling

Structure is 1 alpha subunit

Question 9

Gastric mucosal H+/K+ ATPase

Answer 9

Location: gastric epithelial cell plasma membrane

2H+/2K+ per ATP

H+ secreted into lumen of stomach

2 alpha and 2 beta subunits

Question 10

Structure of P-type ATPases

Answer 10

N-terminus - 4 TMS domains - give rise to 4 stalk domains

C- terminus - 6 TMS domains - gives rise to 5th stalk domain

Most of pump in cytoplasm

Question 11

What are the 3 regions of P-type ATPases

Answer 11

Region B - Conformational changes in ion binding site

Region C - Phosphorylation site/Nucleotide binding site

Region J - Comprises hinge to allow cytosolic regions to move and interact

Question 12

What do conformational changes induce in cation binding?

Answer 12

Binding of ions at low concentration

Dissociation of ions at high concentration

Question 13

SERCA Ca2+ ATPase

Answer 13

ATP hydrolysis + phosphorylation of aspartate

Head regions move

E1 to E2

Alters affinity

Binding site exposed to lumen of SR

Question 14

CPx pumps

Answer 14

Transport toxic and nutrient metal (Cu, Pb, Zn, Cd)

First identified in Enterococcus (COPA and COPB)

Question 15

What’s Menkes disease

Answer 15

Systemic copper deficiency

Question 16

What’s Wilsons disease

Answer 16

Excessive copper accumulation in liver

Question 17

CPx vs P-type pump

Answer 17

N-terminus - 6 TMS domains, cysteine repeats

x in CPx can be serine, histidine, or cysteine

C-region: conserved histidine-proline

C-terminus - less TMS domains (2)

Question 18

V-type pumps

Answer 18

Almost exclusively H+ ATPases on intracellular membranes of eukaryotic cells

Question 19

Explain rotational catalysis

Answer 19

ATP hydrolysis by A3B3 complex

Torque generated in D-subunit

Ring of 6 ‘c’ subunits rotates

H+ translocation through V-pumps

Question 20

ATP binding cassettes (ABC)

Answer 20

Superfamily of transporters

Energy from ATP used to transport solutes in and out

Question 21

Where are ABC superfamily important

Answer 21

Cystic fibrosis

MDR transporters - pathogen and cancer cell drug resistance

Question 22

Structure of ABC transporters

Answer 22

N-terminus - 6 TMS domains

C-terminus - 6 TMS domains

Each 6 joined by ATP

Question 23

ABC conserved motif

Answer 23

GX(S/T)GXGK(S/T)(S/T)

Question 24

Explain flippase model of ABC transporters

Answer 24

Lipid soluble molecule diffuses into cytosolic facing leaflet of plasma membrane

Binds MDR1 protein

Substrate flipped by ATP hydrolysis to exoplasmic leaflet of plasma membrane

Substrate molecule diffuses into extracellular space