Master of Science

thesisMajor Histocompatibility Complex Class II molecules (MHCII) play a critical role in immunity. Of key importance is the presentation of antigenic peptides to CD4+ effector T cells of the adaptive immune system. During the assembly of MHCII molecules, a conserved chaperone protein known as invariant chain binds MHCII with a segment buried in the MHCII binding groove. Invariant chain functions to track MHCII into endocytic peptide loading compartments, prevents inappropriate peptide binding, and assists to maintain the structure of MHCII. After the complex enters the peptide loading compartment invariant chain is sequentially cleaved to leave only a small fragment, class II associated invariant chain (CLIP), bound through the peptide binding groove. CLIP must be removed prior to antigenic peptide loading in the peptide binding groove. The process of peptide exchange occurs through a series of events that are not completely understood. The research described herein is aimed at further elucidated the role that a second peptide plays in the dissociation of a prebound peptide. As with most assays involving the dissociation of a species which could possibly rebind, the peptide dissociation from MHCII has traditionally been performed using an excess of unlabeled peptide which can rapidly block the binding site to prevent rebinding of the labeled peptide. As with most „competition‟ type assays, it was assumed that the unlabeled peptide did not affect the dissociation of bound, labeled peptide. Recent iv evidence has suggested that the unlabeled peptide might play a role in the dissociation of the labeled peptide; in fact this recent data suggests that a second peptide in solution is required for peptide to dissociate from MHCII. By using several different analytical techniques to measure the dissociation in the absence and presence of competitors with varying affinity for MHCII we show that each individual peptide has a rate of dissociation, which occurs independently of the presence of a second peptide in solution. These findings lead to the conclusion that dissociation and exchange of peptides on MHCII is a random process that is determined solely by the affinity of the bound peptide for MHCII