HLA-DM and HLA-DO: bi-directional regulation of function, tuned by pH (APP3P.103)

• DOI: 10.4049/jimmunol.192.supp.111.4
• Publication Date: 2023-01-01T09:09:26Z
• Authors: Wei Jiang; Michael Strohman; Nan Wang; Tieying Hou; Elizabeth Mellins
• Keywords: 0301 basic medicine; 03 medical and health sciences

Abstract Peptide processing and loading onto MHC-II in endosomes occurs across a wide pH range. We studied effects on DM/DO regulation of peptide vitro, using soluble, recombinant molecules, including stabilized mutant DO dimer (sDOAP11A) that inhibits DM normally. tested different DO:DM ratios at 4.6-7.2 during 4 HLA alleles. alone had no effect. inhibition was diminished lower ratios, implying stable association is required for Inhibition occurred the range, but reduced 4.6. Using narrower gradient (4.56-5.30), which function equivalent, we identified as primary target effects. activity after pre-incubation 5.4, longer exposure, higher temperature enhanced this FRET analysis low consistent with release rather than conformational change intact complexes. SPR, bio-layer inferometry ELISA assays showed pH≤4.85, suggesting released binds back inefficiently. With 4.6 or DM-DOwt complexes acquired DM-like activity; active SEC fractions indicated presence free DM. Pre-incubation sDM these pHs caused aggregation. Complexes full-length DOwt similar behavior to soluble molecules. conclude generates by releasing DO, both chaperones