Structural basis of Kbm8 alloreactivity. Amino acid substitutions on the β-pleated floor of the antigen recognition site

H. D. Hunt, J. K. Pullen, R. F. Dick, J. A. Bluestone, L. R. Pease

Research output: Contribution to journalArticlepeer-review

29 Scopus citations


We have analyzed the functional significance of the four amino acid differences between the parental H-2Kb and mutant H-2Kbm8 glycoproteins. Six bm8 variants including single substitutions at residues 22, 23, 24, and 30 as well as paired substitutions at residues 23, 30 and 22, 24 were generated and transfected into L cells. Surface expression of these H-2Kb variants was analyzed using monoclonal antibodies which bind to well-defined H-2Kb epitopes. No alterations introduced into the conformational structure of H-2Kb by the amino acid substitutions were detected. The effect of these substitutions on CTL recognition was initially analyzed using the following bulk CTL: either H-2Kb anti-H-2Kbm8, H-2Kbm8 anti-H-2Kb, or third party anti-H-2Kb. The alloreactivity between H-2Kb and H-2Kbm8 is dominated by the amino acid substitution at residue 24 (Glu→Ser). The complete bm8 phenotype, however, also requires the additional substituion at residue 22 (Tyr→Phe). The H-2Kbm8 anti-Kb bulk CTL reacted with both variant H-2Kbm8 molecules containing single substitutions at amino acid positions 22 or 24 but not the variant molecule containing both substitutions. Further analysis using three individual H-2Kbm8 anti-Kb CTL clones indicated the complexity of the self Kbm8 phenotype. Clone 8B1.20 did not react to changes in residues 22 or 24. The 8B1.32 clone reacted with the change at residue 22 but not with the change at residue 24, although the 8B1.54 clone reacted with the change at residue 24 but not with the change at residue 22. The changes in residues 23 (Met→Ile) and/or 30 (Asp→Asn) did not impact significantly on the alloantigenic properties of Kbm8 as determined by both the bulk and cloned CTL populations. According to the three-dimensional class I structure the substitution at amino acid 24 is inaccessible to the TCR. The location of this substitution within the Ag recognition site implies that altered peptide binding and not a disruption of MHC residues that interact with the TCR, is responsible for the alloreactivity between H-2Kb and H-2Kbm8.

Original languageEnglish (US)
Pages (from-to)1456-1462
Number of pages7
JournalJournal of Immunology
Issue number5
StatePublished - 1990

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology


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