Transcript Slide 1

Lecture 16
Membrane Transport
Active transport
Where would you find active transport?
•interface with the environment….
•maintain cell volume
•control internal environment
•signaling….Ca++ gradient
Characteristics of a Transporter
•Saturability…characterized by KM and Vmax
•Stereospecificity..or specificity unrelared to biophysical
characteristics
•Higher rate than expected from oil/water partition coef.
GLUT = sugar transporters
GLUT1-GLUT12
Michaelis-Menten equation for enzyme/transport
reactions is very similar to the Langmuir isotherm
V  Vmax
[ s]
K m  [ s]
Vmax 1
Km = 1 mM
0.8
Km = 10 mM
0.6
0.4
0.2
0
0
10
20
30
[s], mM
40
50
A “simple explanation” says
that the rate of reaction should
be proportional to the
occupancy of the binding site
as long as Vmax is constant.
Bacterial Lac permease (lacY): Lactose-proton co-transporter
from Abramson et al. 2003
The Lac permease functional cycle,
an example of coupled transport
Note: the proton is always taken up first, but is released at last, which
ensures strict coupling of transport without H+ leakage
from Abramson et al. 2003
[ S ]1



RT
ln(
)  z S F
energy in gradient:
s
[ S ]2
Example:
Na+-glucose symport: stoichiometry of 2:1
at equilibrium: Δμglu= -2ΔμNa
[ Na  ]out
[ glu]in
2RT ln(
)  2F  RT ln(
)

[ glu]out
[ Na ]in
[ Na  ]out
[ glu]in
2F
2 log(
)




log(
)

2.3RT
[ glu]out
[ Na ]in
Aspartate Transporter:
Na+ - dependent
transport of aspartate
(from Boudker et al., Nature
2007)
apical
Tight
junction
K+
Na+ GLU
Na+
G LU
Na+
Na-K ATPase = the
primary active transport,
generates concentration
gradients of Na+ and K+
utilizing ATP
Na-Glucose co-transporter,
utilizes Na+ gradient as a
secondary energy source
Na+
K+
K+
GLU
GLUT
GLU
H2O
Na+
basolateral
Glucose diffusion facilitator
(no energy consumed,
passive transport)
ATPases that couple splitting of ATP with ion motion across the membrane
ATP synthase
(works in reverse)
pump only protons
During contraction of the striated and cardiac muscle, Ca2+ is
released into the cytoplasm, but during the relaxation phase it is
actively pumped back into SR. Ca2+ ATPae (SERCA) constitutes
>80% of total integral protein in SR.
Muscle Ca2+ pump (SERCA)
High-affinity state
open inside
Low-affinity state
open outside
The activity of SERCA, especially in the heart is regulated
by Phospholamban, a small (single-pass) transmembrane
protein. Phosphorylation of phospholamban by PkA
removes its inhibitory action and increases the activity of
SERCA by an order of magnitude.
The activity of plasma membrane Ca2+ pump (p-class) is
regulated by calmodulin, which acts as a sensor of Ca
concentration. Elevated Ca2+ binds to calmodulin, which
in turn causes allosteric activation of the Ca2+ pump.
Post-Alberts Cycle for the Na+/K+ ATPase
Vacuilar or Lysosomal V-type ATPases work in conjunction with Cl- channels
at equilibrium:
 H 
 ATP  n H 
[ H  ]1
 RT ln(  )  F  RTpH  F
[ H ]2
BtuCD ATPase pumps vitamin B12 (ABC transporter)
Many ABC transporters work as flppases or pump lipid-soluble substances (MDR)
MDR1
flippase