A Structurally Altered d,l-Amino Acid TCRα Transmembrane Peptide Interacts with the TCRα and Inhibits T-Cell Activation in Vitro and in an Animal Model

Abstract

Protein-protein interactions in the membrane are pivotal for the cellular response to receptor-sensed stimuli. Recently, it has been demonstrated that an all-d-amino acids analogue of the TCRalpha transmembrane peptide (CP) is recruited to the TCR complex and inhibits T-cell activation in vitro and in vivo, similarly to the wild-type CP peptide. Here we investigated the relative contributions of the secondary structure of CP compared to its side chains in the association of CP with the TCR. We disrupted the secondary structure of CP by replacing two positive residues, needed for the interaction of CP with the TCR complex, by their d-enantiomers (2D-CP). Structure disruption was demonstrated by CD and FTIR spectroscopy, and molecular dynamics simulation in a bilayer environment. In vitro, 2D-CP colocalized with the TCR (visualized with confocal microscopy), immunoprecipitated with TCR but not MHC I, and inhibited T-cell activation. The peptide was effective also in vivo: it inhibited adjuvant arthritis in rats and delayed type hypersensitivity in BALB/c mice. Moreover, 2D-CP manifested greater immunosuppressive activity than wild-type CP, both in vivo and in vitro, which can be attributed to the greater solubility and resistance to degradation of 2D-CP. In molecular terms, these findings suggest that, under certain conditions, protein-protein interactions within the membrane might be more dependent on side chain interactions than on a specific secondary structure. The new altered secondary structure probably determines how the Lys and the Arg are positioned with respect to each other, so they can interact with the TM domain of the receptor. In clinical terms, the increased solubility and resistance to degradation of d-stereoisomers might be exploited in the targeted inactivation of pathogenic signaling pathways such as those arising from TCR-triggered activation of T-cells in immune-mediated disorders.

Single-spanning transmembrane domains in cell growth and cell-cell interactions

“…93 It is also interesting to note that some derivatives of TM peptides, containing D-amino acids or modified with hydrophobic moieties, are also active. [104][105][106][107] These examples all demonstrate that it is possible to design small peptides or peptidomimetics that specifically modulate much larger target membrane proteins by acting within cell membranes. While several questions related to the stability, biodistribution or toxicity of such TM peptides have yet to be addressed, these developments represent the first steps towards a new generation of peptide drugs.…”

Lessons from Viruses: Controlling the Function of Transmembrane Proteins by Interfering Transmembrane Helices

“…D-amino acid containing antimicrobial peptides have been designed [132], and a Damino acid containing peptide, which interacts with the HIV-1 fusion peptide, successfully inhibited the fusion process [133]. Recently, a TM peptide has been synthesized with two D-amino acids that interacts with the human T-cell receptor alpha (TCR-) [134]. While the secondary structure of the D-amino acid containing peptide was disturbed, the interaction with the TCR-was unaffected.…”

Molecular modelling of co-receptor CD8αα and its complex with MHC class I and T-cell receptor in sea bream (Sparus aurata)

“…Structural studies could help in improving our knowledge on the behaviour of these molecules until specific markers for immunoregulatory peptides will be available. Moreover, it may be possible to design some peptides, based on the amino acids present at the interface of the modelled complexes, that can block complex formation and that, therefore, should inhibit T-cell activation in vitro and permit interesting functional analyses, as already performed in mammals [42]. …”