Structural Characterization of Non-Peptide Antigens

Structural Characterization of Non-Peptide Antigens

MHC encoded proteins on antigen presenting cells (APC) bind and present peptide antigen resulting in specific T cell activation. Recently, non-peptide lipid antigens have been shown to be presented to T-cells by human CD1 proteins, a family of nonpolymorphic glycoproteins encoded outside of the MHC and expressed in association with b-2 microglobulin on the surface of antigen presenting cells. Three classes of CD1b-restricted lipid antigens – free mycolic acids, glycosylated mycolates (GMM) and phosphoglycolipids such as phosphatidylinositol mannosides (PIM) or lipoarabinomannan (LAM)(1,2,3) – have been identified. We present here the structure of the first CD1c restricted lipid antigen. Three separate CD1c-restricted human T cell lines were found to recognize three distinct lipids from M. tuberculosis, M. avium or M. smegmatis. The molecular structure of one of these lipid antigens was determined in detail using chemical and mass spectrometric analysis. The antigen was identified as a phospholipid based on its sensitivity to HF treatment (under conditions that hydrolyze phospholipids but not representative lipids or glycolipids), chromatographic behavior and detection of inorganic phosphate by tandem mass spectrometric analysis. The detailed structure was elucidated by electrospray ionization mass spectrometry (ESI-MS) and ESI based collision induced dissociation (CID) and shown to be a hexose phospholipid. Treatment of the CD1c-restricted hexose phospholipid with mild acid resulted in liberation of the hexose and loss of antigenicity, contrasting with CD1b lipids, which require mild acid conditions for their presentation by CD1b. Thus, this CD1c presented antigenic structure is different from all known CD1b presented glycolipids and suggests that CD1c functions in a different, but complementary way to CD1b. 1. E.M. Beckman et al., Nature 360, 593 (1992) 2. P.A. Sieling et al., Science 269, 227 (1995) 3. D.B. Moody et al., ibid. 278, 283 (1997)

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