Objective To clone, express and purify OmpA protein of Acinetobacter baumannii, so as to provide a theoretical basis for further preparation of Ab_OmpA polyclonal antibody and explore the expression of Ab_OmpA in Acinetobacter baumannii and its specific biological activity.

Methods The OmpA gene sequence of AB5075 strain was downloaded from NCBI official website and a pair of specific primers containing the enzyme restriction sites of BamH I and Xho I were synthesized. The OmpA gene was amplified by PCR and the prokaryotic expression recombinant plasmid Ab_OmpA-pET28a was constructed by genetic engineering. Enzyme digestion and sequencing verified the successful connection. The recombinant plasmid Ab_OmpA-pET28a was transformed into E. coli BL21 (DE3) strain, and the expression of OmpA protein was induced by IPTG, and then the OmpA protein was purified by agarice magnetic beads using His label protein. SDS-PAGE adhesive was used to detect the purification effect of OmpA protein, and explore the optimal induction conditions and preserve the protein. ExPASy-Translate Tool, CDD, TMHMM-2.0, IEDB, Resource, ProtParam, ProtScale, SignalP-4.1, PSORT Ⅱ, SOPMA, SWISS-MODEL were used for OmpA biological information analysis.

Results The expected OmpA gene of about 1 500 bp was obtained and successfully cloned into pET-28a plasmid, and the OmpA protein was successfully purified through IPTG induced expression. Bioanalysis of OmpA protein showed that: Acinetobacter Baumannii AB5075_OmpA gene encoding 471 amino acids, protein molecular formula is C₂₁₅₆H₃₅₁₀N₅₉₄O₆₉₃S₄, theoretical protein molecular weight is 48 900, protein isoelectric point is 5.59, instability index is 33.33. The aliphatic index was 100.81 and the mean hydrophilic value was –0.009, making it a hydrophilic stable protein. There were two transmembrane regions but no signaling peptides, and the sequence highly conserved. The secondary structure is mainly Random coil and α helix. The OmpA protein is subcellular to the nucleus and has 12 potential B-cell epitopes. It can interact with multiple proteins.

Conclusions The prokaryotic expression product of OmpA gene in Acinetobacter baumannii was successfully obtained, and the optimal induction condition was determined to be 0.5mmol/L IPTG at 16 ℃ for 24 hours. The preparation of Ab_OmpA polyclonal antibody provides a basis for the detection of nosocomial Acinetobacter baumannii, and the further study of OmpA pathogenic mechanism lays a foundation. OmpA protein has potential immunogenicity and can be used as candidate molecules for OmpA vaccine and drug target research.