Ligand-modulation of the stability of the glucose transporter GLUT 1

The glucose transporter GLUT 1 was isolated from human erythrocytes and rec onstituted into endogenous membrane lipids. Results from thermal denaturati on studies, using differential scanning calorimetry, indicate that the ther mal denaturation temperature of GLUT 1 is significantly lower in the presen ce of ATP. The lowering of this transition temperature is very dependent on pH. At more acidic pH, ATP has a greater effect of lowering the thermal de naturation temperature of the protein. For example, with 4.8 mM ATP, the de naturation endotherm is lowered by over 10 degrees at pH 4.3, whereas at pH 7.4, ATP does not alter this transition temperature. However, a change in pH alone, in the absence of ATP, has very little effect on the denaturation temperature. Both glucose and salt partially reverse the lowering of the t emperature of thermal denaturation caused by ATP. Studies of acrylamide que nching of the Trp residues of GLUT 1 indicate that at neutral pH, ATP incre ases the Stern-Volmer quenching constant, while glucose lowers it. The resu lts indicate that ATP binds to GLUT 1 and destabilizes the native structure , leading to a lowering of the thermal denaturation temperature and an incr ease in acrylamide quenching, The effects of ATP are reversed in part by gl ucose and are also partly electrostatic in nature.