[Prokaryotic Expression and Bioinformatic Analysis of Rv3432c From ].

OBJECTIVE

To express the protein enconded by the 3432 gene of (.) by prokaryotic expression, to analyze the structure of the Rv3432c protein by using bioinformatics software, and to explore for new drug targets against ..

METHODS

The 3432 gene was amplified by PCR using the genomic DNA of the inactivated . strain H37Rv as the template and a recombinant plasmid was constructed with the expression vector pET-28a. The expression products were analyzed by SDS-PAGE and purified using affinity chromatography. The biological properties of Rv3432c were analyzed with Protparam, the Pfam online tool, SOMPA, Protscale, TMHMM Signalp 6.0, NetPhos3.1, SUMOsp 2.0, and SWISS-MODEL.

RESULTS

pET-28a-3432 recombinant plasmid sequencing results were fully consistent with those of the target gene. SDS-PAGE analysis showed that the fusion protein existed in the form of a soluble protein with a relative molecular mass of about 55×10, which matched the expected size. ProtParam analysis showed that the Rv3432c protein was hydrophilic (showing a GRAVY value of －0.079). Rv3432c was a protein with no transmembrane structural domains or signal peptide. The secondary structure of Rv3432c mainly consisted of random coils (39.78%) and α-helix (39.57%) and was relatively loosely structured.

CONCLUSION

We successfully constructed a prokaryotic expression plasmid of the Rv3432c protein and analyzed its structure using bioinformatics, laying the foundation for further research on the role of Rv3432c in the pathogenesis and progression of tuberculosis as well as the identification of new drug targets against ..