Expression of a Recombinant Anti-HIV and Anti-Tumor Protein, MAP30, in Nicotiana tobacum Hairy Roots: A pH-Stable and Thermophilic Antimicrobial Protein

Abstract

In contrast to conventional antibiotics, which microorganisms can readily evade, it is nearly impossible for a microbial strain that is sensitive to antimicrobial proteins to convert to a resistant strain. Therefore, antimicrobial proteins and peptides that are promising alternative candidates for the control of bacterial infections are under investigation. The MAP30 protein of Momordica charantia is a valuable type I ribosome-inactivating protein (RIP) with anti-HIV and anti-tumor activities. Whereas the antimicrobial activity of some type I RIPs has been confirmed, less attention has been paid to the antimicrobial activity of MAP30 produced in a stable, easily handled, and extremely cost-effective protein-expression system. rMAP30-KDEL was expressed in Nicotiana tobacum hairy roots, and its effect on different microorganisms was investigated. Analysis of the extracted total proteins of transgenic hairy roots showed that rMAP30-KDEL was expressed effectively and that this protein exhibited significant antibacterial activity in a dose-dependent manner. rMAP30-KDEL also possessed thermal and pH stability. Bioinformatic analysis of MAP30 and other RIPs regarding their conserved motifs, amino-acid contents, charge, aliphatic index, GRAVY value, and secondary structures demonstrated that these factors accounted for their thermophilicity. Therefore, RIPs such as MAP30 and its derived peptides might have promising applications as food preservatives, and their analysis might provide useful insights into designing clinically applicable antibiotic agents.

Hevein-Like Antimicrobial Peptides Wamps: Structure–Function Relationship in Antifungal Activity and Sensitization of Plant Pathogenic Fungi to Tebuconazole by WAMP-2-Derived Peptides

“…The aliphatic index defined as the relative volume of the side chains of Val, Ala, Ile and Leu proposed by Ikai [42] positively correlates with thermal stability of a protein. For thermophilic proteins it makes about 100 [43]. The aliphatic index of WAMP-2-derived peptides is not high and varies from 6.67 to 32.5 in WAMP-G2, WAMP-N and WAMP-G1, and for WAMP-C it is equal to 0 because of the absence of the above-mentioned amino acids in the peptide ( Table 2).…”

Recombinant expression of LFchimera antimicrobial peptide in a plant-based expression system and its antimicrobial activity against clinical and phytopathogenic bacteria

“…ER's targeting of AMPs can be utilized to promote the accumulation of recombinant AMPs without significantly influencing their biological activity [72]. In another study, the recombinant MAP30 antimicrobial protein was effectively expressed and targeted to ER in Nicotiana tabacum hairy roots; it exhibited significant antibacterial activity [73], and ER-signal was used for successful expression of a plant defencin protein in transgenic tobacco seeds [53]. However, because AMPs have natural destructive effects on microorganisms and are relatively sensitive to proteolytic degradation, they are often produced by fusion to a partner in heterologous hosts and/or by modifying different parameters during fermentation to obtain successful recombinant production [74][75][76].…”

Targeting microbial pathogens by expression of new recombinant dermaseptin peptides in tobacco

“…The use of genetic engineering strategies to introduce genes encoding natural and synthetic antimicrobial peptides is a new approach in engineering resistance to a broad spectrum of pathogens (Zasloff, ). Various recombinant proteins and peptides have so far been introduced in HRs of different plant species, and their antimicrobial properties have been tested in vitro (Aleinein, Schäfer, & Wink, ; Moghadam, Niazi, Afsharifar, & Taghavi, ; Pham, Schäfer, & Wink, ). For instance, an anti‐HIV protein and an anti‐tumor protein MAP30, including ribosome‐inhibiting proteins were produced in tobacco HRs (Moghadam et al, ).…”

“…Various recombinant proteins and peptides have so far been introduced in HRs of different plant species, and their antimicrobial properties have been tested in vitro (Aleinein, Schäfer, & Wink, ; Moghadam, Niazi, Afsharifar, & Taghavi, ; Pham, Schäfer, & Wink, ). For instance, an anti‐HIV protein and an anti‐tumor protein MAP30, including ribosome‐inhibiting proteins were produced in tobacco HRs (Moghadam et al, ). Analysis of total proteins extracted from transgenic HRs indicated a strong antimicrobial activity against both gram‐positive and gram‐negative bacteria as well as pathogenic fungi (Moghadam et al, ).…”

“…For instance, an anti‐HIV protein and an anti‐tumor protein MAP30, including ribosome‐inhibiting proteins were produced in tobacco HRs (Moghadam et al, ). Analysis of total proteins extracted from transgenic HRs indicated a strong antimicrobial activity against both gram‐positive and gram‐negative bacteria as well as pathogenic fungi (Moghadam et al, ). Chahardoli, Fazeli, and Ghabooli () expressed a lactoferricin encoding protein in tobacco HR culture with potent antibacterial activity against Escherichia coli (Chahardoli et al, ).…”