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Secondary structure composition and pH-dependent conformational changes of soluble recombinant HLA-DM
1Department of Pediatrics, Stanford University, Stanford, California 94305, USA. rbushch@leland.stanford.edu
The Journal of Biological Chemistry
|October 9, 1998
Summary
Human Leukocyte Antigen-DM (HLA-DM) undergoes a pH-dependent conformational change, impacting its function in Major Histocompatibility Complex (MHC) class II peptide editing. This structural shift may explain HLA-DM
Area of Science:
- Immunology
- Structural Biology
- Protein Biochemistry
Background:
- Human Leukocyte Antigen-DM (HLA-DM) is crucial for Major Histocompatibility Complex (MHC) class II antigen presentation.
- HLA-DM catalyzes invariant chain release, stabilizes empty MHC class II molecules, and edits peptides.
- The pH-dependent activity of HLA-DM, optimal at pH 4.5-5.5, suggests a role for conformational changes, but the structural basis is unknown.
Purpose of the Study:
- To investigate the secondary structure of HLA-DM and its resemblance to MHC folds.
- To determine if HLA-DM undergoes conformational changes between neutral (pH 7) and endosomal (pH 5) pH conditions.
- To explore the structural basis for the pH-dependent activity of HLA-DM.
Main Methods:
- Recombinant soluble HLA-DM (sDM) was analyzed using far-UV circular dichroism (CD) spectroscopy.
- Fluorescence emission spectra were recorded at different pH values.
- Guanidinium chloride unfolding experiments were performed to assess protein stability.
Main Results:
- Far-UV CD spectra indicate HLA-DM possesses an alpha/beta protein structure, similar to MHC class I and II molecules.
- CD spectra revealed at least three distinct conformational states of sDM across different pH levels, including a pH-dependent shift between neutral and endosomal pH.
- Fluorescence spectroscopy and unfolding experiments showed a slight blue-shift and reduced stability at pH 5 compared to pH 7, indicating a pH-induced conformational change affecting the hydrophobic core and hydrogen bonding.
Conclusions:
- Soluble HLA-DM undergoes a significant pH-dependent conformational change between neutral and endosomal pH.
- This conformational shift involves alterations in hydrogen bonding and the hydrophobic core of HLA-DM.
- The observed structural changes likely contribute to the pH-dependent functional activity of HLA-DM in MHC class II peptide editing.
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