2.4. Bioinformatic Analysis

Based on the gene-cloning results, the amino acid sequence of SkChi65 was deduced using ORF Finder (https://www.ncbi.nlm.nih.gov/orffinder/, accessed on 20 December 2023). The Conserved Domain Database (http://www.ncbi.nlm.nih.gov/Structure/cdd/wrpsb.cgi, accessed on 22 December 2023) was used to predict the typical domains. Secondary structure prediction was performed using SMART (https://smart.embl.de/, accessed on 22 December 2023), multiple sequence alignment was performed using DNAMAN 9.0, and the phylogenetic tree of SkChi65 with the most similar homologs was constructed using MEGA 11.0 by the neighbor-joining method [33]. Analysis and visualization of tertiary structures were performed using Alphafold2 (https://colab.research.google.com/github/sokrypton/ColabFold/blob/main/AlphaFold2.ipynb#scrollTo=kOblAo-xetgx, accessed on 25 March 2024) and PyMOL3.11.