Genetic, enzymatic, and pathogenic studies of the iron superoxide dismutase of Campylobacter jejuni

Abstract

Campylobacter jejuni is a microaerobic bacterium that produces an acute, self-limiting, watery or bloody diarrhea in humans. Little is known about how C. jejuni causes disease or even what specific capabilities it requires for survival in vivo. The enzyme, superoxide dismutase (SOD), which catalyzes the breakdown of superoxide radicals to hydrogen peroxide and dioxygen is one of the bacterial cell's major defense mechanisms against oxidative damage. A PCR-based search for sod genes in C. jejuni 81-176 revealed that this bacterium contained at least one sod gene. We cloned and sequenced a sod gene from 81-176 and determined that its predicted protein product was most similar to that of FeSODs (sodB genes). Transcriptional analysis indicated that this gene is monocistronic and may be transcribed from a o70-like promoter. Nondenaturing polyacrylamide gels stained to reveal SOD activities, accompanied by inhibition studies, demonstrated that C. jejuni produces five electrophoretically distinct bands of SOD activity, all of which appeared to be FeSODs. Analysis of an 81-176 sodB strain revealed that all of these FeSOD activities may be products of the one sodB gene that we cloned. The expression and enzymatic activity of the respective sodB and FeSOD produced by both C. jejuni and Helicobacterpylori were examined in Escherichia coli. Both genes were expressed in E. coli, and the proteins produced were enzymatically active. Finally, the ability of the 81-176 sodB strain to survive INT407 cell invasion was found to be significantly decreased (12-fold) compared with that of the parent, suggesting a potential role for SodB in C. jejuni intracellular survival.

The role of iron in Campylobacter gene regulation, metabolism and oxidative stress defense

“…In contrast, Campylobacter is not likely to grow outside a host, since it is exposed to thermal, osmotic and oxidative stresses. Iron seems to play an important role in both processes; Campylobacters have developed iron binding and transport systems which allow them to acquire su⁄cient iron for growth [15], whereas iron compounds also function as cofactors for the oxidative stress defense systems allowing aerobic survival [9,10,19,75,76,79]. Furthermore, there may be a link between hemin and enterotoxin activity of C. jejuni [111], although the issue of enterotoxin production by C. jejuni is controversial [20].…”

“…The SodB protein is located in the cytoplasm, and was characterized in both C. jejuni and C. coli. C. jejuni and C. coli sodB mutants did not exhibit growth retardation in laboratory conditions, but were attenuated in intracellular survival in epithelial cells [75] and in chicken colonization [10]. The C. coli sodB mutant was also severely a¡ected in its survival in milk and on chicken skin, aerobic survival during stationary phase, and interestingly also freeze^thaw stress [10,11].…”

“…Campylobacter expresses a single species of SOD, which is cofactored by iron (Cj0169, SodB) [75,76]. The SodB protein is located in the cytoplasm, and was characterized in both C. jejuni and C. coli.…”

Role of environmental survival in transmission ofCampylobacter jejuni

“…If the cell is unable to neutralise these toxic compounds, they can lead to protein, nucleic acid and membrane damage. Exposure of Campylobacter to oxygen induces catalase, not superoxide dismutase (SOD), the major defence against oxidative stress in most bacteria (Garenaux et al, 2008), though basal activity of SOD may be important (Pesci et al, 1994). The best described catalase in C. jejuni is encoded for by katA (Cj1385 in C. jejuni NCTC11168) (Day et al, 2000;Atack & Kelly, 2009).…”

Helicobacter pylori: an invading microorganism? A review

“…H. pylori has a superoxide dismutase (SOD) and sequencing and alignment revealed striking homology to the following facultative intracellular human pathogens : Listeria ivanovii, Listeria monocytogenes, Coxiella burnetii, Porphyromonas gingivalis, Legionella pneumophila and Entamoeba histolytica [71]. The H. pylori SOD gene further resembles a SOD gene found in Campylobacter jejuni and it has been shown that the absence of the SOD gene impairs the ability of C. jejuni to survive within epithelial cells (INT407) (12-fold reduction in viability of intracellular organisms) [72]. A similar mechanism could be suspected in H. pylori.…”