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The D0 domain of KIR3D acts as a major histocompatibility complex class I binding enhancer.

Khakoo SI, Geller R, Shin S, Jenkins JA, Parham P - J. Exp. Med. (2002)

Bottom Line: In contrast point mutations in the D1 and D2 domains of KIR3DL1, made from knowledge of KIR2D:HLA-C interactions, disrupted binding to Bw4(+) HLA-B.The results are consistent with a model in which D1 and D2 make the principal contacts between KIR3DL1 and HLA-B while D0 acts through a different mechanism to enhance the interaction.This modulatory role for D0 is compatible with natural loss of expression of the D0 domain, a repeated event in the evolution of functional KIR genes.

View Article: PubMed Central - PubMed

Affiliation: Department of Structural Biology, Stanford University School of Medicine, CA 94305, USA.

ABSTRACT
In contrast to the KIR2D:HLA-C interaction, little is known of KIR3DL1's interaction with HLA-B or the role of D0, the domain not present in KIR2D. Differences in the strength and specificity for major histocompatibility complex class I of KIR3DL1 and its common chimpanzee homologue Pt-KIR3DL1/2 were exploited to address these questions. Domain-swap, deletion, and site-directed mutants of KIR3DL1 were analyzed for HLA-B binding using a novel, positively signaling cell-cell binding assay. Natural 'deletion' of residues 50 and 51 from its D0 domain causes Pt-KIR3DL1/2 to bind Bw4(+) HLA-B allotypes more avidly than does KIR3DL1. Deletion of these residues from KIR3DL1, or their substitution for alanine, enhanced binding of Bw4(+) HLA-B. None of 15 different point mutations in D0 abrogated KIR3DL1 binding to Bw4(+) HLA-B. In contrast point mutations in the D1 and D2 domains of KIR3DL1, made from knowledge of KIR2D:HLA-C interactions, disrupted binding to Bw4(+) HLA-B. The results are consistent with a model in which D1 and D2 make the principal contacts between KIR3DL1 and HLA-B while D0 acts through a different mechanism to enhance the interaction. This modulatory role for D0 is compatible with natural loss of expression of the D0 domain, a repeated event in the evolution of functional KIR genes.

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Naturally occurring variations in D0 stabilize the KIR-MHC class I interaction. (A) Comparison between the HLA-B specificities of KIR3DL1ζ, Pt-KIR3DL1/2ζ, and KIR3DL1-Δ50,51,P14Sζ in the SEAP assay. Jurkat transfectants stably expressing these molecules were incubated with 721.221 transfectants expressing HLA-B*3801, B*1513, B*1502, and B*5801, or untransfected 721.221 cells. In B a further comparison is made with KIR3DL1ζ, Pt-KIR3DL1/2ζ, and KIR3DL1-Δ50,51ζ in the SEAP assay against the same panel of cell lines. The data shown are the means and standard errors of three independent experiments in comparison to plate-bound anti-CD3.
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fig5: Naturally occurring variations in D0 stabilize the KIR-MHC class I interaction. (A) Comparison between the HLA-B specificities of KIR3DL1ζ, Pt-KIR3DL1/2ζ, and KIR3DL1-Δ50,51,P14Sζ in the SEAP assay. Jurkat transfectants stably expressing these molecules were incubated with 721.221 transfectants expressing HLA-B*3801, B*1513, B*1502, and B*5801, or untransfected 721.221 cells. In B a further comparison is made with KIR3DL1ζ, Pt-KIR3DL1/2ζ, and KIR3DL1-Δ50,51ζ in the SEAP assay against the same panel of cell lines. The data shown are the means and standard errors of three independent experiments in comparison to plate-bound anti-CD3.

Mentions: Features unique to the D0 domain of Pt-KIR3DL1/2 are deletion of what are residues 50 and 51 in KIR3DL1 (Fig. 4) and substitution of proline for serine at position 14. To determine the contribution these differences make to the enhanced binding due to the Pt-KIR3DL1/2 D0 domain, a mutant KIR3DL1ζ was made that lacks residues 50 and 51 and has serine substituted for proline at position 14. This Δ50,51,P14Sζ mutant gave a pattern of HLA-B response like that of CHHζ, in that there was enhanced reactivity with the Bw4+ HLA-B allotypes but no reactivity with HLA-B*1502 (Fig. 5 A). Thus, the enhanced response appears specific for allotypes reactive with the wild-type KIR3DL1 molecule. A mutant having just deletion of residues 50 and 51 (Δ50,51ζ) was then made and shown to have the same binding phenotype as Δ50,51,P14Sζ (Fig. 5 B). These results demonstrate that absence of residues 50 and 51 is responsible for the enhanced binding of Bw4+ HLA-B by Pt-KIR3DL1/2 compared with KIR3DL1.


The D0 domain of KIR3D acts as a major histocompatibility complex class I binding enhancer.

Khakoo SI, Geller R, Shin S, Jenkins JA, Parham P - J. Exp. Med. (2002)

Naturally occurring variations in D0 stabilize the KIR-MHC class I interaction. (A) Comparison between the HLA-B specificities of KIR3DL1ζ, Pt-KIR3DL1/2ζ, and KIR3DL1-Δ50,51,P14Sζ in the SEAP assay. Jurkat transfectants stably expressing these molecules were incubated with 721.221 transfectants expressing HLA-B*3801, B*1513, B*1502, and B*5801, or untransfected 721.221 cells. In B a further comparison is made with KIR3DL1ζ, Pt-KIR3DL1/2ζ, and KIR3DL1-Δ50,51ζ in the SEAP assay against the same panel of cell lines. The data shown are the means and standard errors of three independent experiments in comparison to plate-bound anti-CD3.
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Related In: Results  -  Collection

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fig5: Naturally occurring variations in D0 stabilize the KIR-MHC class I interaction. (A) Comparison between the HLA-B specificities of KIR3DL1ζ, Pt-KIR3DL1/2ζ, and KIR3DL1-Δ50,51,P14Sζ in the SEAP assay. Jurkat transfectants stably expressing these molecules were incubated with 721.221 transfectants expressing HLA-B*3801, B*1513, B*1502, and B*5801, or untransfected 721.221 cells. In B a further comparison is made with KIR3DL1ζ, Pt-KIR3DL1/2ζ, and KIR3DL1-Δ50,51ζ in the SEAP assay against the same panel of cell lines. The data shown are the means and standard errors of three independent experiments in comparison to plate-bound anti-CD3.
Mentions: Features unique to the D0 domain of Pt-KIR3DL1/2 are deletion of what are residues 50 and 51 in KIR3DL1 (Fig. 4) and substitution of proline for serine at position 14. To determine the contribution these differences make to the enhanced binding due to the Pt-KIR3DL1/2 D0 domain, a mutant KIR3DL1ζ was made that lacks residues 50 and 51 and has serine substituted for proline at position 14. This Δ50,51,P14Sζ mutant gave a pattern of HLA-B response like that of CHHζ, in that there was enhanced reactivity with the Bw4+ HLA-B allotypes but no reactivity with HLA-B*1502 (Fig. 5 A). Thus, the enhanced response appears specific for allotypes reactive with the wild-type KIR3DL1 molecule. A mutant having just deletion of residues 50 and 51 (Δ50,51ζ) was then made and shown to have the same binding phenotype as Δ50,51,P14Sζ (Fig. 5 B). These results demonstrate that absence of residues 50 and 51 is responsible for the enhanced binding of Bw4+ HLA-B by Pt-KIR3DL1/2 compared with KIR3DL1.

Bottom Line: In contrast point mutations in the D1 and D2 domains of KIR3DL1, made from knowledge of KIR2D:HLA-C interactions, disrupted binding to Bw4(+) HLA-B.The results are consistent with a model in which D1 and D2 make the principal contacts between KIR3DL1 and HLA-B while D0 acts through a different mechanism to enhance the interaction.This modulatory role for D0 is compatible with natural loss of expression of the D0 domain, a repeated event in the evolution of functional KIR genes.

View Article: PubMed Central - PubMed

Affiliation: Department of Structural Biology, Stanford University School of Medicine, CA 94305, USA.

ABSTRACT
In contrast to the KIR2D:HLA-C interaction, little is known of KIR3DL1's interaction with HLA-B or the role of D0, the domain not present in KIR2D. Differences in the strength and specificity for major histocompatibility complex class I of KIR3DL1 and its common chimpanzee homologue Pt-KIR3DL1/2 were exploited to address these questions. Domain-swap, deletion, and site-directed mutants of KIR3DL1 were analyzed for HLA-B binding using a novel, positively signaling cell-cell binding assay. Natural 'deletion' of residues 50 and 51 from its D0 domain causes Pt-KIR3DL1/2 to bind Bw4(+) HLA-B allotypes more avidly than does KIR3DL1. Deletion of these residues from KIR3DL1, or their substitution for alanine, enhanced binding of Bw4(+) HLA-B. None of 15 different point mutations in D0 abrogated KIR3DL1 binding to Bw4(+) HLA-B. In contrast point mutations in the D1 and D2 domains of KIR3DL1, made from knowledge of KIR2D:HLA-C interactions, disrupted binding to Bw4(+) HLA-B. The results are consistent with a model in which D1 and D2 make the principal contacts between KIR3DL1 and HLA-B while D0 acts through a different mechanism to enhance the interaction. This modulatory role for D0 is compatible with natural loss of expression of the D0 domain, a repeated event in the evolution of functional KIR genes.

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