Development and characterization of a catalytically inactive cysteine protease domain of RtxA1/MARTX_Vv as a potential vaccine for Vibrio vulnificus

Abstract

Recent studies have defined several virulence factors as vaccine candidates against Vibrio vulnificus. However, most of these factors have the potential to cause pathogenic effects in the vaccinees or induce incomplete protection. To overcome these drawbacks, a catalytically inactive form, CPD Vv (C3725S), of the well-conserved cysteine protease domain (CPD) of V. vulnificus multifunctional autoprocessing repeats-in-toxin (MARTX Vv /RtxA1) was recombinantly generated and characterized. Notably, active and passive immunization with CPD Vv (C3725S) conferred protective immunity against V. vulnificus strains. These results may provide a novel framework for developing safe and efficient subunit vaccines and/or therapeutics against V. vulnificus that target the CPD of MARTX toxins.Key words cysteine protease domain, protective immunity, RtxA1/MARTX Vv , Vibrio vulnificus.Infection with Vibrio vulnificus, an important opportunistic human pathogen, is associated with seafood consumption and exposure of wounds to seawater containing the bacteria (1-4). Despite aggressive antibiotic therapy and supportive care, these infections frequently progress to severe skin lesions and septicemia with a mortality rate exceeding 50% (3,(5)(6)(7)(8). The annual incidence of V. vulnificus infection has been increasing worldwide, including in the USA, Japan, Korea and Taiwan (6,[9][10][11][12][13]. However, no vaccine to control this lethal bacterial disease has yet been licensed for human use.Although recent studies have shown that traditionally inactivated V. vulnificus cells and several virulence determinants such as V. vulnificus capsular polysaccharide, quorum sensing-dependent metalloprotease and multifunctional autoprocessing repeats-in-toxin (MARTX Vv or RtxA1) can induce protective immunity against V. vulnificus infection (14-17), these antigens for possible vaccine development also have the potential to elicit cytotoxicity or induce incomplete protection, both of which could complicate vaccine design (18-23). For example, V. vulnificus RtxA1/MARTX Vv , a large secreted MARTX family protein, has several effector domains, including extensive repeats at both the N and C termini, an actin cross-linking domain, a Rho inactivation domain and a CPD (24-27). These repeats and domains are associated with multiple cytotoxic and cytopathic activities (28)(29)(30)(31)(32)(33)(34). Collectively, these observations imply that successful development of a highly effective V. vulnificus vaccine depends on designing conserved immunogens in diverse V. vulnificus strains, thus both avoiding potential risk(s) and eliciting broad protective immunity.Of the potential vaccine targets for V. vulnificus, the C-terminal region (amino acids 3491-4701; molecular weight %130 kDa) of V. vulnificus RtxA1/MARTX Vv , RtxA1-C, harbors highly conserved regions including CPD and GD-rich repeats (14,(24)(25)(26)(35)(36)(37)(38)(39) a recent study suggested that the C-terminal %130 kDa cleavage product of secreted RtxA1/MARTX Vv from V. vulnificus might be ...

Cross‐protection against Vibrio cholerae infection by monoclonal antibodies against Vibrio vulnificus RtxA1/MARTXVv

“…Recent studies have shown that protective immunity against Vibrio species is induced by both bacterial components and toxins, including LPS, pili, capsular polysaccharides, cholera toxins and MARTX toxins . Among these protective immunogens, MARTX toxins, which comprise a subfamily of large secreted RTX proteins, have been identified in a number of Vibrio species and other pathogenic gram‐negative bacteria .…”

Immunogenicity and protective efficacy of <i>Vibrio vulnificus</i> flagellin protein FlaB in a wound infection model

“…Improvement of this rFlaB vaccine will be needed in the future. Flagellin has already been found to have high adjuvant activity [ 19 , 24 ], and therefore should be combined with another additional antigen such as Vibrio vulnificus hemolysin VVH, metalloprotease VvpE, and/or RTX toxin [ 3 , 10 , 13 , 14 , 16 ]. There have been many reports on utilization of these proteins as vaccine antigens, and immunization with these proteins confers effective protection against lethal challenge by V. vulnificus on hosts [ 10 ].…”