Thermal stability of MHC class I-β₂-microglobulin peptide complexes in the endoplasmic reticulum is determined by the peptide occupancy of the transporter associated with antigen processing complex

Once MHC class I heavy chain binds β₂-microglobulin (β₂m) within the endoplasmic reticulum, an assembly complex comprising the class I heterodimer, TAP, TAPasin, calreticulin, and possibly Erp57 is formed before the binding of high affinity peptide. TAP-dependent delivery of high affinity peptide to in vitro translated K^bβ₂m complexes within microsomes (TAP⁺/TAPasin⁺) was studied to determine at which point peptide binding becomes resistant to thermal denaturation. It was determined that the thermal stability of K^b-β₂m-peptide complexes depends on the timing of peptide binding to K^bβ₂m relative to TAP binding high affinity peptide. Premature exposure of the TAP complex to high affinity peptide before its association with class I heavy chain results in K^bβ₂m-peptide-TAP complexes that lose peptide upon exposure to elevated temperature after solubilization away from microsome-associated proteins. These findings suggest that the order in which class I heavy chain associates with endoplasmic reticulum-resident chaperones and peptide determines the stability of K^bβ₂m-peptide complexes.