Class I major histocompatibility complex (MHC) heterodimer, composed of human leukocyte antigen (HLA)-A2 heavy chain and human β₂-microglobulin (β₂m), was produced by denaturation and gel filtration of the recombinant water-soluble HLA-A2/β₂m/peptide ternary complex in 8 M urea Tris−HCl buffer, followed by refolding of the separated chains without peptide. Peptide affinity and kinetics of the ternary complex formation and dissociation were investigated in real time by monitoring the fluorescence resonance energy transfer (FRET) from intrinsic HLA-A2 heavy-chain tryptophans to a dansyl fluorophore conjugated to the bound peptide. Peptide binding to the heterodimer was a second order process with rate constants linearly dependent upon temperature in Arrhenius coordinates over 0−20 °C. The binding rate constant of pRT6C-dansyl [ILKEPC(dansyl)HGV] at 37 °C evaluated by extrapolation of the Arrhenius plot was (2.0 ± 0.5) × 10⁶ M^-1 s^-1. Association of the heavy chain with β₂m was a first order process, apparently controlled by a conformational transition in the heavy chain. One of these conformations bound to β₂m to form the heavy chain/β₂m heterodimer whereas the second conformer oligomerized. Peptide dissociation from the ternary complex was a first-order reaction over the temperature range 20−37 °C, suggesting that the ternary complex also exists in two conformations. Taken together, the present data suggest that association of β₂m changes the HLA-A2 heavy-chain conformation thereby promoting peptide binding. Peptide dissociation from the ternary complex induces dissociation of the heavy-chain/β₂m heterodimer thereby causing oligomerization of the heavy chain. The lability of the HLA-A2/β₂m heterodimer and the strong tendency of the “free” heavy chain to oligomerize may provide an efficient mechanism for control of antigen presentation under physiological conditions by reducing the direct loading of HLA with exogenous peptide at the cell surface.