Enzymatic and antibacterial activity of the recombinant endolysin PVP-SE1gp146 expressed in Hansenula polymorpha

Antibiotic resistance, a major global concern, highlights the need for discovering alternative therapies. Recently, endolysins have garnered attention as antibacterial tools with a lower resistance development rate compared to conventional antibiotics, and their production in various expression hosts holds significance. Given its generally recognized as safe (GRAS) status and other advantages, Hansenula polymorpha offers a promising host for endolysin production. PVP-SE1gp146 originates from the Salmonella Enteritidis-specific phage PVP-SE1, which has been previously characterized. We inserted the PVP-SE1gp146 coding gene into the H. polymorpha expression vector pHIPX4. The resulting recombinant, pHIPX4-PVP-SE1gp146, was then introduced into H. polymorpha NCYC495 to facilitate the production of the endolysin PVP-SE1gp146. The expression level of the PVP-SE1gp146 protein was assessed, and it was determined to be approximately 43 mg/l of yeast culture medium. The enzymatic (muralytic) activity of this endolysin was also evaluated, corresponding to the version produced by the E. coli Bl21 strain. The endolysin exhibited admissible antibacterial activity against several gram-negative species, including P. aeruginosa, E. coli, and A. baumannii, while showing an almost negligible impact on K. pneumoniae. Endolysin production within GRAS-approved hosts holds potential for combating antibiotic-resistant bacteria. Challenges involve optimizing concentrations, targeting gram-negative species and improving attachment to bacterial cell walls. Addressing these issues requires dedicated research in endolysin engineering and a comprehensive evaluation of their production in diverse expression hosts.