Isolation and Characterization of Three Porin‐Like Proteins from Vibrio vulnificus: Effect of Different Growth Media on Their Production

Abstract

The effect of the culture media on the composition of the outer membrane protein of Vibrio vulnificus strain 393 from human blood was examined. Only one major outer membrane protein, with an apparent molecular weight of 37,000 (37K protein) and 34,000 (34K protein), was formed in the cells grown in 3% NaCl-BHI broth and chemically defined medium, respectively. The production of one major outer membrane protein was also observed in other isolates from humans and asari clam when they were grown in 3% NaCl-BHI broth. On the other hand, three major outer membrane proteins, with apparent molecular weights of 48,000 (48K protein), 37,000 (37K protein), and 34,000 (34K protein), were produced in the cells grown in 3% NaCl-nutrient broth. Three proteins, 48K, 37K, and 34K from strain 393, were purified and the amino acid compositions were determined. Although there was a little difference in the composition of amino acid among three proteins, the amino acid compositions of the three porin-like proteins showed characteristic properties of the porins of Escherichia coli and Salmonella typhimurium. Immunoblot analysis of the outer membrane proteins from four vibrios, E. coli, and S. typhimurium using monospecific antisera against these three porin-like proteins showed that only the antiserum against 37K protein cross-reacted with the outer membrane proteins from all the strains tested.Key words: Porin-like protein, Vibrio vulnificus Vibrio vulnificus, a halophilic marine bacterium, causes a number of extraintestinal infections. Septicemia caused by V. vulnificus is associated with underlying diseases including hemochromatosis, thalassemia, chronic cirrhosis and hepatoma (2). In view of the high correlation between V. vulnificus infections and diseases involving increased iron status, the effect of iron and iron-chelating agents upon the pathogenicity has been examined (18). In regard to the outer membrane, there are several papers describing the iron-regulated outer membrane proteins which are produced under the ironlimited conditions. However, the number of manuscript describing a porin which plays an essential role in the uptake of nutrition is limited. In a previous paper (9), we reported that the composition of outer membrane protein was affected by the composition of the media used for cultivation. In the present study, we cultured V. vulnificus strain 393 grown in 3% NaCl-brain heart infusion (BHI) broth, which are often used to cultivate the bacteria, and compared the composition of the outer membrane proteins with those of cells cultured in 3% NaCl-nutrient broth and in chemically defined medium. It became clear that the strain produces a new major outer membrane protein, the 37K protein, when it is grown in 3% NaCl-BHI broth. We also examined the outer membrane proteins from other V. vulnificus strains isolated from the blood of patients with septicemia, from seawater and from an asari clam grown in 3% NaCl-BHI broth.As strain 393 produces three porin-like proteins when they are cultured in 3% NaCl-nutri...

Utilization of hemin and hemoglobin by Vibrio vulnificus biotype 2

“…This finding suggests that the heme prosthetic group of Hb is the one recognized by the Hb-binding protein(s). Recently, proteins of similar molecular mass in biotype 1 have been identified as porin-like proteins (24). Similar results have been found with the fish pathogen V. anguillarum, which has Hm-and Hb-binding proteins with very similar molecular masses (33).…”

Molecular Basis of Bacterial Outer Membrane Permeability Revisited

“…We do not yet know, however, the properties of OmpL and OmpH that might make them suitable for environments with different pressures. Vibrio vulnificus was shown to express, depending on the growth medium, a few porin-like proteins (345); with the recent sequencing of the genome of this species (GenBank accession numbers NC004459 and NC004460), a more detailed study may appear in the future. Vibrio cholerae is known to produce two porins: OmpU (120), whose transcription is activated by the regulatory protein ToxR (155), and OmpT, which is repressed by ToxR (374).…”

Acid adaptation induces cross-protection against some environmental stresses in Vibrio parahaemolyticus.

“…Cell envelope pellets were prepared from acid-adapted and nonadapted cells as described previously (Koga and Takumi, 1994). The cell envelope preparation was then treated with sodium lauroyl sarcosinate (Sarkosyl), as described previously (Koga and Takumi, 1995b).…”