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
Vmax 1 Km = 1 mM 0.8 Km = 10 mM 0.6 0.4 0.2 0 0 10 20 30 40 50 [s], mM Michaelis-Menten equation for enzyme/transport reactions is very similar to the Langmuir isotherm 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
energy in gradient: Example: Na+-glucose symport: stoichiometry of 2:1 at equilibrium: Δμglu= -2ΔμNa
Aspartate Transporter: Na+ - dependent transport of aspartate (from Boudker et al., Nature 2007)
apical Na-K ATPase = the primary active transport, generates concentration gradients of Na+ and K+ utilizing ATP Tight junction Na-Glucose co-transporter, utilizes Na+ gradient as a secondary energy source GLUT Glucose diffusion facilitator (no energy consumed, passive transport) H2O basolateral
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.
Vacuilar or Lysosomal V-type ATPases work in conjunction with Cl- channels at equilibrium:
Many ABC transporters work as flppases or pump lipid-soluble substances (MDR) MDR1 flippase