Rezaei Roya, Moazamian Elham, Montazeri-Najafabady Nima
Department of Microbiology, College of Sciences, Agriculture and Modern Technology, Shiraz Branch, Islamic Azad University, Shiraz, Iran.
Shiraz Endocrine and Metabolism Research Center, Shiraz University of Medical Sciences, Shiraz, Iran.
Mol Biol Rep. 2023 May;50(5):4469-4480. doi: 10.1007/s11033-023-08378-5. Epub 2023 Apr 4.
Parasporin (PS) proteins have cytocidal activity preferential for various human malignant cells. The purpose of this investigation was to see if the PS separated from B. thuringiensis strain E8 isolate had any particular cytotoxicity against breast cancer.
The extracted spores-crystal proteins were solubilized and digested with proteinase K. Cytotoxicity effects were analysed by MTT assay. Caspases activities were measured using ELISA. SDS-PAGE analysis was performed for determination of molecular weight of Cry protein. Identification of extracted proteins function was evaluated by MALDI-TOF MS analysis. Breast cancer cells line (MCF-7) was highly susceptible to 1 mg/mL PS and showed apoptosis characteristics, but it has no effects on the normal cells (HEK293). Apoptosis evaluation showed that caspases 1, 3, 9 and BAX were remarkably up-regulated in cancer cells, indicating the intrinsic pathway activation in these cells. PS Size was determined using SDS-PAGE in E8 isolate as 34 kDa and a 25 kDa digested peptide was identified as PS4. The function of PS4 was reported as an ABC-transporter by spectrometry.
The data of the present study show that PS4 is a selective cytotoxic protein against breast cancer and a molecule with a lot of potentials for next researches.
伴胞晶体蛋白（PS）对多种人类恶性细胞具有优先的杀细胞活性。本研究的目的是观察从苏云金芽孢杆菌菌株E8分离物中分离出的PS对乳腺癌是否具有任何特殊的细胞毒性。
将提取的芽孢晶体蛋白溶解并用蛋白酶K消化。通过MTT法分析细胞毒性作用。使用ELISA测定半胱天冬酶活性。进行SDS-PAGE分析以确定Cry蛋白的分子量。通过MALDI-TOF MS分析评估提取的蛋白质功能的鉴定。乳腺癌细胞系（MCF-7）对1mg/mL的PS高度敏感并表现出凋亡特征，但对正常细胞（HEK293）没有影响。凋亡评估显示癌细胞中的半胱天冬酶1、3、9和BAX显著上调，表明这些细胞中的内源性途径被激活。在E8分离物中使用SDS-PAGE测定PS大小为34kDa，并且鉴定出一个25kDa的消化肽为PS4。通过光谱法报道PS4的功能为ABC转运蛋白。
本研究的数据表明PS4是一种针对乳腺癌的选择性细胞毒性蛋白，并且是一种在后续研究中具有很大潜力的分子。
