[Preparation and activity validation of PP7 bacteriophage-like particles displaying PAP114-128peptide].
Objective To obtain the PP7 bacteriophage-like particles carrying the peptide of prostatic acid phosphatase PAP114-128, and prove that they retain the original biological activity. Methods First, the plasmid pETDuet-2PP7 was constructed as follows: the gene of PP7 coat protein dimer was amplified by gene mutation combined with overlapping PCR technology, and inserted into the vector pETDuet-1. Following that, the plasmid pETDuet-2PP7-PAP114-128was constructed as follows: the PP7 coat protein gene carrying the coding gene of PAP114-128peptide was amplified using PCR, and then inserted into the vector pETDuet-2PP7. Both pETDuet-2PP7 and pETDuet-2PP7-PAP114-128were transformed into E.coli and expressed. The expression product was verified by SDS-PAGE, double immunodiffusion assay and ELISA. Results The gene fragment of PP7 coat protein dimer was obtained by overlapping PCR using Ex Taq DNA polymerase, and the antigenicity of its expression product was the same as that of the coat protein of wild-type PP7 bacteriophage. Moreover, the PAP114-128peptide epitope that was displayed on the surface of PP7 bacteriophage was identical with the corresponding epitope of natural human PAP, and it was able to induce high levels of antibodies. Conclusion The gene of PP7 coat protein dimer with repeated sequences can be prepared by gene mutation combined with overlapping PCR. Based on this, PP7 bacteriophage-like particles carrying PAP peptide can be prepared, which not only solves the problem of the instability of the peptides, but also lays a foundation for the study on their delivery and function.
Department of Laboratory Medicine, Weifang Medical University, Weifang 261053, China.
Yanli
Y
Sun
Department of Hematology, Weifang People's Hospital, Weifang 261000, China. *Corresponding author, E-mail: zhgzbxforget@163.com.
Yanhua
Y
Sun
D000135:Acid Phosphatase / Q000235:genetics* / Q000276:immunology; D000595:Amino Acid Sequence; D000818:Animals; D001435:Bacteriophages / Q000235:genetics* / Q000378:metabolism; D019281:Dimerization; D015870:Gene Expression; D006801:Humans; D007114:Immunization; D051379:Mice; D008810:Mice, Inbred C57BL; D010455:Peptides / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D010957:Plasmids / Q000235:genetics / Q000378:metabolism
D010455:Peptides; D000135:Acid Phosphatase; C428439:prostatic acid phosphatase