Structural analyses of Cryptosporidium parvum epitopes reveal a novel scheme of decapeptide binding to H-2Kb

Yongli Wang; Yankai Chang; Fangyuan Yin; Chunliu Kang; Yao Meng; Fukang Xu; Yiran Liu; Yunxia Zhang; Changjing Wu; Shuhua Fan; Junlong Zhao

doi:10.1016/j.jsb.2025.108168

Structural analyses of Cryptosporidium parvum epitopes reveal a novel scheme of decapeptide binding to H-2K^b

J Struct Biol. 2025 Jan 12:108168. doi: 10.1016/j.jsb.2025.108168. Online ahead of print.

Authors

Yongli Wang¹, Yankai Chang², Fangyuan Yin³, Chunliu Kang³, Yao Meng³, Fukang Xu³, Yiran Liu³, Yunxia Zhang³, Changjing Wu³, Shuhua Fan⁴, Junlong Zhao⁵

Affiliations

¹ State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, PR China; College of Life Science and Agronomy, Zhoukou Normal University, Zhoukou, PR China.
² College of Veterinary Medicine, Henan Agricultural University, Zhengzhou, PR China.
³ College of Life Science and Agronomy, Zhoukou Normal University, Zhoukou, PR China; Institute of Neuroscience and Translational Medicine, Zhoukou Normal University, Zhoukou, PR China.
⁴ College of Life Science and Agronomy, Zhoukou Normal University, Zhoukou, PR China; Institute of Neuroscience and Translational Medicine, Zhoukou Normal University, Zhoukou, PR China. Electronic address: fanshuhuayan@126.com.
⁵ State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, PR China. Electronic address: Zhaojunlong@mail.hzau.edu.cn.

PMID: 39809366
DOI: 10.1016/j.jsb.2025.108168

Abstract

Cryptosporidium has gained much attention as a major cause of diarrhea worldwide. Here, we present the first structure of H-2K^b complexed with a decapeptide from Cryptosporidium parvum Gp40/15 protein (Gp40/15-VTF10). In contrast to all published structures, the aromatic residue P3-Phe of Gp40/15-VTF10 is anchored in pocket C rather than the canonical Y/F at P5 or P6 reported for octapeptides and nonapeptides. The results of in vitro refolding assays and circular dichroism experiments showed that the side chains of P3 and P5 play key roles in Gp40/15-VTF10 peptide binding. However, functional analysis of decapeptide epitopes revealed that the Gp40/15-VTF10 peptide did not elicit a strong CD8⁺T immune response, whereas the decapeptide epitope MEDLE2-INF10 induced a significant CD8⁺ T-cell response in peptide-immunized C57BL/6 mice. Using a model structure of H-2K^b-INF10 complex, we found that the antigenic decapeptide INF10 exhibits a completely different conformation, with the aromatic anchors P3F and P7F docked into the D and C pockets, respectively, while similar peptide conformation and hydrogen bond interactions between the peptide and major histocompatibility complex were found in the resolved H-2K^b-SVF9 complex. As the H-2K^b molecule predominantly prefers octapeptides with a strong anchor of P5 Y/F (or P6 Y/F for nonapeptides) binding to the C pocket, we propose that P7 Y/F in the C pocket may represent a novel binding mode for decapeptides. The results should increase the accuracy of T-cell epitope prediction and support the development of T-cell epitope vaccines against cryptosporidiosis.

Keywords: Cell-mediated immunity; Cryptosporidium parvum; Decapeptide epitope; H-2K(b).