Recombinant Production of a Novel Fusion Protein: Listeriolysin O Fragment Fused to S1 Subunit of Pertussis Toxin. 2021

Hossein Forghani, and Mahin Jamshidi Makiani, and Hossein Zarei Jaliani, and Mina Boustanshenas, and Seyed Mohsen Zahraei
Antimicrobial Resistance Research Center, Institute of Immunology and Infectious Diseases, Iran University of Medical Sciences, Tehran, Iran.

Some resources have suggested that genetically inactivated pertussis toxoid (PTs) bear a more protective effect than chemically inactivated products. This study aimed to produce new version of PT, by cloning an inactive pertussis toxin S1 subunit (PTS1) in a fusion form with N-terminal half of the listeriolysin O (LLO) pore-forming toxin. Deposited pdb structure file of the PT was used to model an extra disulfide bond. Codon-optimized ORF of the PTS1 was used to make recombinant constructs of PTS1 and LLO-PTS1 in the pPSG-IBA35 vector. The recombinant PTS1 and LLO-PTS1 proteins were expressed in BL21 DE3 and SHuffle T7 strains of E. coli and purified by affinity chromatography. Cytotoxic effects of the recombinant proteins were examined in the MCF-7 cell line. The purity of the products proved to be more than 85%, and the efficiency of the disulfide bond formation in SHuffle T7 strain was higher than BL21 DE3 strain. No cytotoxicity of the recombinant proteins was observed in MCF-7 cells. Soluble recombinant PTS1 and LLO-PTS1 proteins were produced in SHuffle T7 strain of E. coli with high efficiency of disulfide bonds formation. The LLO-PTS1 with corrected disulfide bonds was successfully expressed in E. coli SHuffle T7 strain. Due to the safety for human cells, this chimeric molecule can be an option to prevent pertussis disease if its immunostimulatory effects would be confirmed in the future.

UI MeSH Term Description Entries
D010567 Pertussis Vaccine A suspension of killed Bordetella pertussis organisms, used for immunization against pertussis (WHOOPING COUGH). It is generally used in a mixture with diphtheria and tetanus toxoids (DTP). There is an acellular pertussis vaccine prepared from the purified antigenic components of Bordetella pertussis, which causes fewer adverse reactions than whole-cell vaccine and, like the whole-cell vaccine, is generally used in a mixture with diphtheria and tetanus toxoids. (From Dorland, 28th ed) Vaccine, Pertussis
D011993 Recombinant Fusion Proteins Recombinant proteins produced by the GENETIC TRANSLATION of fused genes formed by the combination of NUCLEIC ACID REGULATORY SEQUENCES of one or more genes with the protein coding sequences of one or more genes. Fusion Proteins, Recombinant,Recombinant Chimeric Protein,Recombinant Fusion Protein,Recombinant Hybrid Protein,Chimeric Proteins, Recombinant,Hybrid Proteins, Recombinant,Recombinant Chimeric Proteins,Recombinant Hybrid Proteins,Chimeric Protein, Recombinant,Fusion Protein, Recombinant,Hybrid Protein, Recombinant,Protein, Recombinant Chimeric,Protein, Recombinant Fusion,Protein, Recombinant Hybrid,Proteins, Recombinant Chimeric,Proteins, Recombinant Fusion,Proteins, Recombinant Hybrid
D004220 Disulfides Chemical groups containing the covalent disulfide bonds -S-S-. The sulfur atoms can be bound to inorganic or organic moieties. Disulfide
D004926 Escherichia coli A species of gram-negative, facultatively anaerobic, rod-shaped bacteria (GRAM-NEGATIVE FACULTATIVELY ANAEROBIC RODS) commonly found in the lower part of the intestine of warm-blooded animals. It is usually nonpathogenic, but some strains are known to produce DIARRHEA and pyogenic infections. Pathogenic strains (virotypes) are classified by their specific pathogenic mechanisms such as toxins (ENTEROTOXIGENIC ESCHERICHIA COLI), etc. Alkalescens-Dispar Group,Bacillus coli,Bacterium coli,Bacterium coli commune,Diffusely Adherent Escherichia coli,E coli,EAggEC,Enteroaggregative Escherichia coli,Enterococcus coli,Diffusely Adherent E. coli,Enteroaggregative E. coli,Enteroinvasive E. coli,Enteroinvasive Escherichia coli
D006360 Heat-Shock Proteins Proteins which are synthesized in eukaryotic organisms and bacteria in response to hyperthermia and other environmental stresses. They increase thermal tolerance and perform functions essential to cell survival under these conditions. Stress Protein,Stress Proteins,Heat-Shock Protein,Heat Shock Protein,Heat Shock Proteins,Protein, Stress
D006460 Hemolysin Proteins Proteins from BACTERIA and FUNGI that are soluble enough to be secreted to target ERYTHROCYTES and insert into the membrane to form beta-barrel pores. Biosynthesis may be regulated by HEMOLYSIN FACTORS. Hemolysin,Hemolysins,Hemalysins,Proteins, Hemolysin
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D001427 Bacterial Toxins Toxic substances formed in or elaborated by bacteria; they are usually proteins with high molecular weight and antigenicity; some are used as antibiotics and some to skin test for the presence of or susceptibility to certain diseases. Bacterial Toxin,Toxins, Bacterial,Toxin, Bacterial
D015202 Protein Engineering Procedures by which protein structure and function are changed or created in vitro by altering existing or synthesizing new structural genes that direct the synthesis of proteins with sought-after properties. Such procedures may include the design of MOLECULAR MODELS of proteins using COMPUTER GRAPHICS or other molecular modeling techniques; site-specific mutagenesis (MUTAGENESIS, SITE-SPECIFIC) of existing genes; and DIRECTED MOLECULAR EVOLUTION techniques to create new genes. Genetic Engineering of Proteins,Genetic Engineering, Protein,Proteins, Genetic Engineering,Engineering, Protein,Engineering, Protein Genetic,Protein Genetic Engineering
D049109 Cell Proliferation All of the processes involved in increasing CELL NUMBER including CELL DIVISION. Cell Growth in Number,Cellular Proliferation,Cell Multiplication,Cell Number Growth,Growth, Cell Number,Multiplication, Cell,Number Growth, Cell,Proliferation, Cell,Proliferation, Cellular

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