A sequential model for peptide binding and transport by the transporters associated with antigen processing

Area of Science:

• Immunology
• Molecular Biology
• Cell Biology

Background:

• Transporter associated with antigen processing (TAP) proteins are crucial for translocating antigenic peptides into the endoplasmic reticulum (ER).
• Investigating TAP specificity is challenging due to TAP-independent factors affecting peptide accumulation in ER transport assays.

Purpose of the Study:

• To develop an overexpression system enabling separate quantification of peptide binding to TAP and peptide transport by TAP.
• To elucidate the factors influencing peptide accumulation in the ER during TAP-mediated transport.

Main Methods:

• Development of an overexpression system to independently measure peptide binding to the TAP substrate-binding site and TAP-mediated peptide transport.
• Analysis of peptide accumulation in the ER in relation to binding affinity, glycosylation interactions, and peptide efflux.

Main Results:

• Peptide accumulation efficiency in the ER correlates with binding affinity to the TAP substrate-binding site.
• Peptide accumulation is further modulated by interactions with the ER glycosylation system and potentially peptide efflux.
• Peptide mixtures of 9-16 amino acids exhibit significantly higher affinity for the TAP binding site compared to shorter or longer peptides.

Conclusions:

• Peptide binding to TAP heteromers occurs in the absence of ATP, with ATP binding facilitating peptide release, proposing a functional model for TAP.
• The developed system allows for a more precise understanding of TAP-mediated peptide translocation and its regulatory mechanisms.