Low CO2 permeability of cholesterol-containing liposomes detected by stopped-flow fluorescence spectroscopy

摘要

Here we ask the following: 1) what is the CO2 permeability (Pco(2)) of unilamellar liposomes composed of l--phosphatidylcholine (PC)/l--phosphatidylserine (PS) = 4:1 and containing cholesterol (Chol) at levels often occurring in biologic membranes (50 mol), and 2) does incorporation of the CO2 channel aquaporin (AQP)1 cause a significant increase in membrane Pco(2)? Presently, a drastic discrepancy exists between the answers to these two questions obtained from mass-spectrometric O-18-exchange measurements (Chol reduces Pco(2) 100-fold, AQP1 increases Pco(2) 10-fold) vs. from stopped-flow approaches observing CO2 uptake (no effects of either Chol or AQP1). A novel theory of CO2 uptake by vesicles predicts that in a stopped-flow apparatus this fast process can only be resolved temporally and interpreted quantitatively, if 1) a very low CO2 partial pressure (pCO(2)) is used (e.g., 18 mmHg), and 2) intravesicular carbonic anhydrase (CA) activity is precisely known. With these prerequisites fulfilled, we find by stopped-flow that 1) Chol-containing vesicles possess a Pco(2) = 0.01cm/s, and Chol-free vesicles exhibit approximate to 1 cm/s, and 2) the Pco(2) of 0.01 cm/s is increased 10-fold by AQP1. Both results agree with previous mass-spectrometric results and thus resolve the apparent discrepancy between the two techniques. We confirm that biologic membranes have an intrinsically low Pco(2) that can be raised when functionally necessary by incorporating protein-gas channels such as AQP1.Tsiavaliaris, G., Itel, F., Hedfalk, K., Al-Samir, S., Meier, W., Gros, G., Endeward, V. Low CO2 permeability of cholesterol-containing liposomes detected by stopped-flow fluorescence spectroscopy.