Biochemical Characteristics Of Eimeria Tenella Histone Deacetylase3(EtHDAC3)

Eimeria spp. are one class of the most important pathogenic agent, and can lead to a tremendous economic loss in poultry industry worldwide. Since1950s, the control of this economically important disease has been mainly dependent on the prophylactic medication in feed, by which has been boosting and ensuring the global development of poultry industry. However, this once effective method has been currently encountering with the resistance against anticoccidials by Eimeria spp. in practice, and the novel drugs/drug-targeted molecules should be discovered for the demand of veterinary practices. It has been confirmed in other parasitic protozoa that histone deacetylases (HDACs) are quite promising drug target. HDAC3is a member of the of class I of HDACs family and an important drug target.In this paper, the ORF sequence of HDAC3of Eimeria tenella (EtHDAC3) predicted by bioinformatics method, and cloned from E. tenella Guangdong strain using the primers designed based on this predicted coding sequence. The length of amplified EtHDAC3is1200bp, which encode a peptide with399amino acid. Stucture and function of EtHDAC3were analyzed by bioinformatics methods.The results showed EtHDAC3has complete conserved domains of Class I of HDACs family, and is highly conserved with other apicomplexan which sequence similarity is around90%.The recombinant plasmid pMAL-C2X-EtHDAC3was transformed into E.coli transetta strain and was expressed under induction of IPTG. The results showed transformed bacteria produced a recombinant protein of87Ku in lysates and best temperature was16℃for this protein production. When the concentration of IPTG changes between0.3mmol/L-1.2mmol/L, IPTG has no effect on the expression of EtHDAC3.Real-time PCR was used to detect the mRNA level of EtHDAC3in E. tenella different life stages. Our results revealed that the highest expression of EtHDAC3mRNA are in the sporulated7h oocysts.The3D structure of EtHDAC3was modeled using Swiss-Model based on homologous alignment, and compared with ChHDAC3. The differences in binding kinetics of EtHDAC3and/or ChHDAC3with small inhibitor, Apicidin, was analysed using molecular docking method with AutoDock4.2software. The comparison of3D structures of ChHDAC3and EtHDAC3showed that both of them is composed of8B-sheets and11a-helices. The amino acids in enzymatic active sites are highly conserved, and only have one amino acid difference in their surface recognition sites. Docking results suggest that based on the predicted binding energy, Apicidin is more patent to EtHDAC3than ChHDAC3, and more tightly bind to the EtHDAC3active sites.