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Open Access Publications from the University of California

Understanding receptor specificity through the massively variable major tropism determinant of Bordetella bacteriophage

  • Author(s): Miller, Jason L.
  • et al.

Only a few protein folds are known to tolerate massive sequence variation for the sake of binding diversity. The immunoglobulin (Ig)-fold found in the structural design of antibodies and T-cell receptors as well as the leucine- rich repeat-fold (LRR's) discovered in the variable lymphocyte receptors of the agnathous hagfish are capable of withstanding recombinant variability on the order of 1014-1016 unique protein sequences. Our studies have elucidated the structure of Bordetella bacteriophage major tropism determinant (Mtd) which utilizes the C-type lectin (CLec) fold to accommodate variation for nearly 1013 sequences. Additionally, we have identified other diversity generating retroelement (DGR) encoded proteins in prokaryotes and bacteriophage that also use the CLec- fold for sequence variation. The structure of Mtd is stabilized as an intertwined, pyramidal shaped trimer consisting of three distinct domains with the variable residues organized as discrete receptor binding sites located near the C-termini. The variable residues are presented on surface exposed loops that are stabilized by a network of hydrogen bonds along the C-Lec-fold resulting in a highly static scaffold for mutagenic variation. Association between Mtd-P1 variant with its native receptor, pertactin ectodomain (Prn-E) was characterized and used as a model for understanding receptor binding specificity. Mtd-P1 was co-crystallized with Prn-E and the structure solved to 3.16 Å resolution. We show that despite a difference in structure fold and interaction modalities, Mtd uses its variable residues for target recognition in an antibody-like manner. In addition, we have identified and characterized an antibody-like affinity maturation process where a non-productive interaction is converted to a productive one. More significantly, we have discovered that phage avidity acts as a differential amplifier for binding affinity and specificity. These results indicate that the C-Lec fold acts as a stable platform for sequence variation with enough flexibility for multiple receptor recognition. Finally, our evidence suggests that multivalency is conserved in conferring efficacy and specificity to sequence variable repertoires

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