关键词: 鲍曼不动杆菌, 外膜蛋白34, 包涵体, 圆二色谱, 凋亡

Abstract: The present study is to clone and purify the outer membrane protein 34（Omp34）from Acinetobacter baumannii ATCC 19606，and to analyze its biological structure and effects on the apoptosis of HEK293 cells. First，the gene of outer membrane protein 34 of A. baumannii ATCC 19606 was amplified by PCR，it was cloned into the pET28a（+）vector to generate the pET28a（+）-Omp34 expression vector，and the vector was transformed into Escherichia coli BL21 for expression. Then，the recombinant protein was expressed by IPTG induction and purified by Ni-affinity chromatography. L-arginine and GSH/GSSG were added in the dialysis buffer to facilitate the Omp34 protein correct folding. The biological structure of Omp34 was predicted by bioinformatics. The secondary structure of the Omp34 was detected by circular dichroism（CD）.The apoptosis of HEK293 cells induced by Omp34 were measured by flow cytometry and Western blot. As results，the pET28a（+）-Omp34 recombinant expression vector was successfully constructed，the Omp34 was highly expressed in prokaryotic expression system，and the soluble protein was obtained by refolding. Bioinformatics predicted that Omp34 was beta barrel-shape. CD confirmed that the secondary structure of purified Omp34 was beta sheets，of which the α-helix，β-sheets，and β-turn，and random coil of the Omp34 were 12%，39%，22%，and 27%，respectively. The re-natured Omp34 was able to induce the apoptosis of HEK293 cells. In summary，the Omp34 of A. baumannii can be expressed by molecular cloning technology. Moreover，its secondary structure was analyzed and it is demonstrated that Omp34 can lead to apoptosis of HEK293 cells. This work provides a basis for further study of the biological functions and antibiotic resistance mechanism of A. baumannii Omp34.

Key words: Acinetobacter baumannii, outer membrane protein 34, inclusion body, circular dichroism, apoptosis