TY - JOUR
T1 - Kinetic studies of cytoplasmic antigen processing and production of MHC class I ligands
AU - Stargardt, Anita
AU - Reits, Eric
PY - 2013
Y1 - 2013
N2 - MHC class I molecules present peptides that are derived from intracellular proteins degraded by proteasomes. These peptides often require additional trimming by peptidases to fit into the peptide-binding grove of MHC class I. However, most peptides are rapidly recycled by the large heterogeneous pool of peptidases. Here, we describe a technique to quantify peptide degradation both in living cells and in cell lysates, using quenched peptides that contain a quencher and fluorophore. As degradation results in separation of the quencher and fluorophore, fluorescence will increase. This technique enables the examination of changes in peptide length and amino acid sequence on its half-life, and hence its chances to become presented by MHC class I
AB - MHC class I molecules present peptides that are derived from intracellular proteins degraded by proteasomes. These peptides often require additional trimming by peptidases to fit into the peptide-binding grove of MHC class I. However, most peptides are rapidly recycled by the large heterogeneous pool of peptidases. Here, we describe a technique to quantify peptide degradation both in living cells and in cell lysates, using quenched peptides that contain a quencher and fluorophore. As degradation results in separation of the quencher and fluorophore, fluorescence will increase. This technique enables the examination of changes in peptide length and amino acid sequence on its half-life, and hence its chances to become presented by MHC class I
U2 - https://doi.org/10.1007/978-1-62703-218-6_4
DO - https://doi.org/10.1007/978-1-62703-218-6_4
M3 - Article
C2 - 23329477
SN - 1064-3745
VL - 960
SP - 41
EP - 51
JO - Methods in molecular biology (Clifton, N.J.)
JF - Methods in molecular biology (Clifton, N.J.)
ER -