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Final
discussion and outlook
5
Final discussion and outlook
As antigen presentation by MHC II molecules plays a pivotal role in the adaptive
immune response it is important to understand how antigenic peptides are selected and
loaded onto MHC II molecules which is catalyzed by DM. The crystal structures of
MHC II and DM are already known since 1993 and 1998, respectively, (Brown et al.,
1993; Mosyak et al., 1998) and mutagenesis studies mapped a large lateral surface on
both molecules required for interaction which has been confirmed by tethering
experiments (Busch et al., 2002; Stratikos et al., 2002). However, the molecular
mechanism of how DM catalyzes peptide exchange on MHC II molecules is still
unknown and different models have been proposed including DM targeting the
hydrogen bond network between peptide and MHC II (Mosyak et al., 1998; Weber et
al., 1996) as well as global conformational changes (Belmares et al., 2002).
Recent studies in the lab showed strong DM binding to a low stability DR/peptide
complex missing three N-terminal peptide residues (Anders et al., 2011) and thereby
disrupting conserved MHC II-peptide interactions. The question arose whether the
truncated peptide represented an intermediate state of a DR/peptide complex with the
peptide N-terminus partially released due to peptide mobility. However, some high-
affinity complexes, e.g. DR1/HA, were believed to be resistant to DM action as no DM-
facilitated peptide exchange had been observed (Kropshofer et al., 1996; Sloan et al.,
1995) arguing against this model which implied DM binding to any MHC II/peptide
complex, even if to a small extent. Crucial surface plasmon resonance experiments
undertaken during this study revealed concentration- and temperature-dependent DM
binding to the high-affinity complexes DR1/HA and DR2/MBP, previously exhibiting
no and only little DM binding, respectively. These data demonstrated that at high
temperature DM binding can be detected to high-affinity MHC II/peptide complexes
previously believed to be DM resistant substantiating the relevance of peptide mobility
for DM catalyzed peptide exchange and supporting the model of spontaneous release of
peptide N-terminus (Anders et al., 2011). Furthermore, the functional significance of the
SPR data was demonstrated during these studies by comparing DM
binding experiments
with DM activity data measured by fluorescence polarization displaying an agreement
between both assays.
NMR experiments performed during this work further addressed peptide mobility in
the peptide-binding groove revealing the presence of multiple peptide conformations for