Biophys J, October 2000, p. 1967-1975, Vol. 79, No. 4

Interaction of the Noncovalent Molecular Adapter, -Cyclodextrin, with the Staphylococcal -Hemolysin Pore

Li-Qun Gu^* and Hagan Bayley^*

 ^*Department of Medical Biochemistry and Genetics, The Texas A & M University System Health Science Center, College Station, Texas 77843-1114, and  Department of Chemistry, Texas A & M University, College Station, Texas 77843-3255 USA

Cyclodextrins act as noncovalent molecular adapters when lodged in the lumen of the -hemolysin (HL) pore. The adapters act as binding sites for channel blockers, thereby offering a basis for the detection of a variety of organic molecules with HL as a biosensor element. To further such studies, it is important to find conditions under which the dwell time of cyclodextrins in the lumen of the pore is extended. Here, we use single-channel recording to explore the pH- and voltage-dependence of the interaction of -cyclodextrin (CD) with HL. CD can access its binding site only from the trans entrance of pores inserted from the cis side of a bilayer. Analysis of the binding kinetics shows that there is a single binding site for CD, with an apparent equilibrium dissociation constant that varies by >100-fold under the conditions explored. The dissociation rate constant for the neutral CD molecule varies with pH and voltage, a result that is incompatible with two states of the HL pore, one of high and the other of low affinity. Rather, the data suggest that the actual equilibrium dissociation constant for the HL · CD complex varies continuously with the transmembrane potential.

Biophys J, October 2000, p. 1967-1975, Vol. 79, No. 4
© 2000 by the Biophysical Society   0006-3495/00/10/1967/09  $2.00

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