The Crystal Structure of the MHC Class I (MHC-I) Molecule in the Green Anole Lizard Demonstrates the Unique MHC-I System in Reptiles

Abstract

The reptile MHC class I (MCH-I) and MHC class II proteins are the key molecules in the immune system; however, their structure has not been investigated. The crystal structure of green anole lizard peptide–MHC-I–β2m (pMHC-I or pAnca-UA*0101) was determined in the current study. Subsequently, the features of pAnca-UA*0101 were analyzed and compared with the characteristics of pMHC-I of four classes of vertebrates. The amino acid sequence identities between Anca-UA*0101 and MHC-I from other species are <50%; however, the differences between the species were reflected in the topological structure. Significant characteristics of pAnca-UA*0101 include a specific flip of ∼88° and an upward shift adjacent to the C terminus of the α1- and α2-helical regions, respectively. Additionally, the lizard MHC-I molecule has an insertion of 2 aa (VE) at positions 55 and 56. The pushing force from 55-56VE triggers the flip of the α1 helix. Mutagenesis experiments confirmed that the 55-56VE insertion in the α1 helix enhances the stability of pAnca-UA*0101. The peptide presentation profile and motif of pAnca-UA*0101 were confirmed. Based on these results, the proteins of three reptile lizard viruses were used for the screening and confirmation of the candidate epitopes. These data enhance our understanding of the systematic differences between five classes of vertebrates at the gene and protein levels, the formation of the pMHC-I complex, and the evolution of the MHC-I system.

Peptidomes and Structures Illustrate How SLA-I Micropolymorphism Influences the Preference of Binding Peptide Length

“…Therefore, there are certain limitations in studying the peptide length preference of MHC-I alleles by MHC-I ligandomes in vivo, which further affects the prediction, screening and identification of long peptide epitopes. RPLD-MS can determine the MHC-I random peptide ligandome in vitro and quickly identify MHC-I restricted peptide epitopes, which is especially suitable for studying unknown animal MHC-I molecules lacking research background and conditions (17,19,(31)(32)(33). Our previous studies proved that RPLD-MS can sensitively reflect the different nonapeptide binding capabilities in vitro between SLA-1*04:01 and SLA-1*13:01 caused by a single residue variation (99Y/F) (19).…”

“…RPLD-MS can determine the MHC-I random peptide ligandome in vitro and quickly identify MHC-I restricted peptide epitopes, which is especially suitable for studying unknown animal MHC-I molecules lacking research background and conditions ( 17 , 19 , 31 – 33 ). Our previous studies proved that RPLD-MS can sensitively reflect the different nonapeptide binding capabilities in vitro between SLA-1*04:01 and SLA-1*13:01 caused by a single residue variation (99Y/F) ( 19 ).…”

“…Finally, the peptide-binding motif of MHC-I was determined. To date, the peptide binding properties of MHC-I molecules of five species, pig, bat, frog, lizard and shark, have been identified by RPLD-MS (17,19,(31)(32)(33). This overcomes the limitations of laboratory animals, cell lines and antibodies, and lowers the threshold for the study of animal MHC-I.…”

“…The interactions between the peptide and the PBG of pSLA-1*13:01 RW12 . Tyr 59 , Glu 62 , Glu 63 , Leu 163 , Glu 166 , Ser 167 , Arg 170 , Tyr 171 Tyr 7 , Phe33 , Tyr 59 , Glu 63 , Tyr 159 , Leu 163 , Ser 167 , Tyr 171 Tyr 9 , Met45 , Glu 63 , Lys 66 , Val 67 , Asn 70 , Phe 99 , Tyr 159 Tyr 74 , Gly 77 , Thr 80 , Lue 81 , Tyr 84 , Leu 95 , Ser 97 , Arg 114 , Asp 116 , Thr 143 , Lys 146 , Trp 147…”

Structure and evolution of the squamate major histocompatibility complex as revealed by two Anolis lizard genomes

“…Hung et al (Hung, 2013) documented high amounts of class I diversity in invasive brown anoles (Anolis sagrei), which could contribute to this species' success in colonizing the southern United States. The crystal structure of the MHC I molecule in Anolis carolinensis was also recently solved and found to be unique among previous structures from other lineages (Wang et al, 2021). However, despite these studies of MHC in anoles and the status of the green anole as the first released squamate reference genome, no focused investigation of the composition and structure of the MHC region has been carried out in the green anole genome.…”