The Journal of Experimental Medicine
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© The Rockefeller University Press, 0022-1007/1998/12/2139/ $5.00
The Journal of Experimental Medicine, Volume 188, Number 11, December 7, 1998 2139-2149

Articles

Alteration of a Single Hydrogen Bond between Class II Molecules and Peptide Results in Rapid Degradation of Class II Molecules after Invariant Chain Removal

Stephanie Ceman*, Shenhong Wu*, Theodore S. Jardetzky§, and Andrea J. Sant*,{ddagger}

From the * Department of Pathology, and {ddagger} Committees on Immunology and Cancer Biology, University of Chicago, Illinois 60637; and the § Department of Biochemistry, Molecular Biology, and Cell Biology, Northwestern University, Evanston, Illinois 60208

To characterize the importance of a highly conserved region of the class II β chain, we introduced an amino acid substitution that is predicted to eliminate a hydrogen bond formed between the class II molecule and peptide. We expressed the mutated β chain with a wild-type {alpha} chain in a murine L cell by gene transfection. The mutant class II molecule (81βH–) assembles normally in the endoplasmic reticulum and transits the Golgi complex. When invariant chain (Ii) is coexpressed with 81βH–, the class II–Ii complex is degraded in the endosomes. Expression of 81βH– in the absence of Ii results in a cell surface expressed molecule that is susceptible to proteolysis, a condition reversed by incubation with a peptide known to associate with 81βH–. We propose that 81βH– is protease sensitive because it is unable to productively associate with most peptides, including classII–associated invariant chain peptides. This model is supported by our data demonstrating protease sensitivity of peptide-free wild-type I-Ad molecules. Collectively, our results suggest both that the hydrogen bonds formed between the class II molecule and peptide are important for the integrity and stability of the complex, and that empty class II molecules are protease sensitive and degraded in endosomes. One function of DM may be to insure continuous groove occupancy of the class II molecule.

Key Words: major histocompatibility complex class II • peptide • hydrogen bond • invariant chain • proteolysis

Address correspondence to Dr. Andrea J. Sant, University of Chicago, Dept. of Pathology and Committee on Immunology, 5841 S. Maryland Ave. MC1089, Chicago, IL 60637. Phone: 773-702-3990; Fax: 773-702-3701; E-mail: asant{at}flowcity.bsd.uchicago.edu

S. Ceman is supported by National Institutes of Health grant F32 AI09218 and the Weber Fellowship Fund. T.S. Jardetzky is supported by the International Frontier Science Program and the Pew Memorial Trust. A.J. Sant is supported by National Institutes of Health grant R01 AI34359.

1 Abbreviations used in this paper: aa, amino acids; CHO, Chinese hamster ovary; CLIP, class II–associated invariant chain peptides; cys, cystatin-C; ER, endoplasmic reticulum; GPI, glycan-phosphatidylinositol; HPAP, human placental alkaline phosphatase; Ii, invariant chain; PK, proteinase K; RT, room temperature; WT, wild-type.


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