RESUMEN
In order to characterise grass carp MHC class I (Ctid-MHC I) sequences, 26 Ctid-MHC I genes were cloned from 12 individuals and their alpha domain lineages were analysed. Simultaneously, a quantitative reverse transcription-polymerase chain reaction (Q-RT-PCR) assay was developed to detect Ctid-MHC I tissue-specific expression. The results suggested that Ctid-MHC I could be divided into eight lineages (Ctid-NA-Ctid-NH). Based on whether they contained the motif of eight key amino acids (YYRTKWYY), Ctid-MHC I lineages were divided into two groups [Ctid-MHC I (8(+)) and Ctid-MHC I (8(-))]. The expression analysis showed that the Ctid-MHC I locus/loci appeared in the kidney, gill, intestine, heart, spleen, liver, and brain. A GenBank homology BLAST was performed independently with each alpha domain, and Ctid-MHC I alpha1, alpha2, and alpha3 were categorised into two (V and IX), five (II, IV-VII), and four (IV-VII) domain lineages, respectively. Based on the alphabetic labelling system created in our earlier studies, one alpha1 (IX), four alpha2 (IV-VII), and unique alpha3 (V-VII) domain lineages were observed in grass carp and across the teleostean species.
Asunto(s)
Carpas/genética , Genes MHC Clase I/genética , Filogenia , Secuencia de Aminoácidos/genética , Animales , Carpas/clasificación , Carpas/inmunología , Clonación Molecular , Cartilla de ADN/química , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/veterinaria , Alineación de Secuencia/veterinariaRESUMEN
No information is available to date on the different allelelic structures of the chicken MHC class I (BF2) and beta2m proteins. To elucidate the structure, new allelic beta2m and five different BF2 genes were expressed solubly and purified in a pMAL-p2X/E. coli TB1 system. The 2D structure was detected by circular dichroism (CD) spectroscopy, and the 3D structures of their peptide-binding domain (PBD) were analyzed by homology modeling. The sequence lengths of the alpha-helix, beta-sheet, turn, and random coil in the five BF2 proteins were 69-73 aa, 67-72 aa, 35-37 aa, and 94-98 aa, respectively. The new beta2m protein displayed a typical beta-sheet. Homology modeling of the different BF2 and beta2m proteins demonstrated similarities to the structure of human and rat MHC class I proteins. The 3D structure, however, revealed that the BF2 and beta2m structures were unique. The correct refolding of recombinant BF2 and beta2m proteins might be a powerful tool to further detect antigenic peptides.