The mechanism of peptide exchange by
MHC class II molecules
Inaugural – Dissertation
to obtain the academic degree
Doctor rerum naturalium (Dr. rer. nat.)
submitted to
the
Department of Biology, Chemistry and Pharmacy of
Freie Universität Berlin
by
Monika-Sarah
Schulze
from Giessen
2012
II
This thesis is based on research conducted from 2008 to 2012 at the Dana-Farber
Cancer Institute in Boston, MA, USA, under the supervision of Prof. Dr. Kai
Wucherpfennig.
Gutachter:
Erster Gutachter: Prof. Dr.
Kai Wucherpfennig
Dana-Farber Cancer Institute
Harvard
Medical School
Zweiter Gutachter: Prof. Dr. Udo Heinemann
Max-Delbrück-Center
for Molecular Medicine
Freie Universität Berlin
Disputationstermin: 15. Mai 2012
V
Summary
Summary
Presentation of antigenic peptides by major histocompatibility complex class II
(MHC II) molecules on the cell surface is essential as it evokes different immune
responses including antibody production, cell destruction, and initiation of regulatory
mechanisms. Peptide loading and formation of stable MHC II/peptide complexes is
catalyzed by the MHC II like protein HLA-DM (DM). A major focus of this work was
to understand the molecular mechanism of DM-mediated peptide exchange which
would aid in efforts to predict immunogenicity of known and emerging pathogens.
Furthermore, enhanced peptide exchange of MHC II molecules by a synthetic small
molecule, J10, was investigated. This small molecule has therapeutic potential as it
could allow actively modifying the presented peptide repertoire which is of interest for
various medicinal applications, e.g. peptide based vaccination. To address the dynamic
process of peptide exchange, which involves breaking and reforming multiple peptide-
MHC II interactions, various methods were applied including X-ray crystallography,
NMR spectroscopy and surface plasmon resonance (SPR).
SPR experiments undertaken during this doctoral work revealed definitive DM
binding at increased temperature to the high-affinity MHC II/peptide complexes
DR1/HA and DR2/MBP previously exhibiting no or only little DM susceptibility. This
finding supports the model of a common transient MHC II/peptide conformation for
high- and low-affinity MHC II/peptide complexes which is dependent on kinetic
parameters and more abundant at higher temperature. Furthermore, the functional
significance of the SPR data was demonstrated by correlating DM binding to a high-
and a low-affinity MHC II/peptide complex with DM activity on the two complexes as
measured
by fluorescence polarization, showing that the two assays were in agreement.
Previous studies showed that release of the peptide N-terminus is crucial for DM
binding and may be facilitated by spontaneous peptide motion which is also supported
by SPR and NMR data obtained during this doctoral work. However, the structural
implications of a partial peptide release were unknown. In this doctoral thesis a DR1
structure is presented carrying an HA peptide variant missing the two N-terminal
peptide residues (P-2, P-1) that represents an intermediate state of a MHC II molecule
during peptide release. Surprisingly, no major but small conformational changes were