Single liposome analysis of peptide translocation by the ABC transporter XXXX .

@entity1 @entity62333 @entity10056 @entity855
@entity855 -binding cassette (ABC) transporters use @entity855 to drive solute transport across biological membranes. Members of this superfamily have crucial roles in cell physiology, and some of the transporters are linked to severe diseases. However, understanding of the transport mechanism, especially of @entity1 ABC exporters, is scarce. We reconstituted the @entity1 lysosomal polypeptide ABC transporter @entity62333 , expressed in @entity10056 , into lipid vesicles (liposomes) and performed explicit transport measurements. We analyzed solute transport at the single liposome level by monitoring the coincident fluorescence of solutes and proteoliposomes in the focal volume of a confocal microscope. We determined a turnover number of eight peptides per minute, which is two orders of magnitude higher than previously estimated from macroscopic measurements. Moreover, we show that @entity62333 translocates peptides against a large concentration gradient. Maximal filling is not limited by an electrochemical gradient but by trans-inhibition. Countertransport and reversibility studies demonstrate that peptide translocation is a strictly unidirectional process. Altogether, these data are included in a refined model of solute transport by ABC exporters.
