CSTA  Vol.4 No.4 , November 2015
Single-Chain Expression and Crystallization of an Antigenic C-Terminus in Complex with the Regulatory Domain of ER Aminopeptidase 1
Abstract: Human endoplasmic reticulum aminopeptidase 1 (ERAP1) is one of two ER luminal aminopeptidases that participate in the final processing of peptide precursors and generates the N-termini of the MHC class I-restricted epitopes. In order to investigate the interactions of its binding site with substrate peptides, X-ray crystallographic analyses have been carried out to study structures of ERAP1 regulatory (ERAP1_R) domain in complex with antigenic peptides. Single-chain bimodular constructs with various antigenic peptides linked to the C-terminal end of ERAP1_R domain are designed to facilitate crystallization process of these complexes. These recombinant proteins have been purified and crystalized, and x-ray diffraction data of one crystal have been processed to a resolution of 2.8 . The crystal belongs to the space group P21, with unit cell parameters a =64.2, b = 66.8, c = 66.3 , β = 110.2°. A Refmac-refined omit map reveals a clear density for the antigenic peptide’s carboxylate-end that is in contact with the ERAP1 regulatory domain of neighboring molecule. Thus the single-chain bimodular constructs have provided an expedited approach to study sequence-specific interactions between the ERAP1 regulatory domain and antigen peptide’s C-terminal ends.
Cite this paper: Sui, L. , Gandhi, A. and Guo, H. (2015) Single-Chain Expression and Crystallization of an Antigenic C-Terminus in Complex with the Regulatory Domain of ER Aminopeptidase 1. Crystal Structure Theory and Applications, 4, 47-52. doi: 10.4236/csta.2015.44006.

[1]   Saric, T., Chang, S.C., Hattori, A., York, I.A., Markant, S., Rock, K.L. and Goldberg, A.L. (2002) An IFN-γ—Induced Aminopeptidase in the ER, ERAP1, Trims Precursors to MHC Class I—Presented Peptides. Nature Immunology, 3, 1169-1176.

[2]   York, I.A., Chang, S.C., Saric, T., Keys, J.A., Favreau, J.M., Goldberg, A.L. and Rock, K.L. (2002) The ER Aminopeptidase ERAP1 Enhances or Limits Antigen Presentation by Trimming Epitopes to 8-9 Residues. Nature Immunology, 3, 1177-1184.

[3]   Chang, S.C., Momburg, F., Bhutani, N. and Goldberg, A.L. (2005) The ER Aminopeptidase, ERAP1, Trims Precursors to Lengths of MHC Class I Peptides by a “Molecular Ruler” Mechanism. Proceedings of the National Academy of Sciences of the United States of America, 102, 17107-17112.

[4]   Guo, H.C., Jardetzky, T.S., Garrettt, T.P., Lane, W.S., Strominger, J.L. and Wiley, D.C. (1992) Different Length Peptides Bind to HLA-Aw68 Similarly at Their Ends But Bulge out in the Middle. Nature, 360, 364-366.

[5]   Fremont, D.H., Matsumura, M., Stura, E.A., Peterson, P.A. and Wilson, I.A. (1992) Crystal Structures of Two Viral Peptides in Complex with Murine MHC Class I H-2Kb. Science, 257, 919-927.

[6]   Kochan, G., Krojer, T., Harvey, D., Fischer, R., Chen, L., Vollmar, M., and Oppermann, U. (2011) Crystal Structures of the Endoplasmic Reticulum Aminopeptidase-1 (ERAP1) Reveal the Molecular Basis for N-Terminal Peptide Trimming. Proceedings of the National Academy of Sciences of the United States of America, 108, 7745-7750.

[7]   Evnouchidou, I., Momburg, F., Papakyriakou, A., Chroni, A., Leondiadis, L., Chang, S.C., and Stratikos, E. (2008) The Internal Sequence of the Peptide-Substrate Determines Its N-Terminus Trimming by ERAP1. PLoS ONE, 3, e3658-e3658.

[8]   Nguyen, T.T., Chang, S.C., Evnouchidou, I., York, I.A., Zikos, C., Rock, K.L. and Stern, L.J. (2011) Structural Basis for Antigenic Peptide Precursor Processing by the Endoplasmic Reticulum Aminopeptidase ERAP1. Nature Structural & Molecular Biology, 18, 604-613.

[9]   Gandhi, A., Lakshminarasimhan, D., Sun, Y. and Guo, H.C. (2011) Structural Insights into the Molecular Ruler Mechanism of the Endoplasmic Reticulum Aminopeptidase ERAP1. Scientific Reports, 1, 1-5.

[10]   R?tzschke, O., Falk, K., Stevanovic, S., Gnau, V., Jung, G. and Rammensee, H.G. (1994) Dominant Aromatic/Ali- phatic C-Terminal Anchor in HLA-B* 2702 and B* 2705 Peptide Motifs. Immunogenetics, 39, 74-77.

[11]   Fleischhauer, K., Avila, D., Vilbois, F., Traversari, C., Bordignon, C. and Wallny, H.J. (1994) Characterization of Natural Peptide Ligands for HLA-B* 4402 and -B* 4403: Implications for Peptide Involvement in Allorecognition of a Single Amino Acid Change in the HLA-B44 Heavy Chain. Tissue Antigens, 44, 311-317.