• Journal of Microbiology
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• v.33 no.4
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• pp.283-288
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• 1995

We have identified hemolysin rendering virulency of Vibrio anguilarum grown at 23.deg.C which was evaluated on human RBCs. Hemolysin itself was separated as a single band on non-denaturing gel electrophoresis. Vibrio hemolysin was destroyed by trypsin and proteinase K and was heat labile. Optimal pH for activity was around pH 6 while pI of the molecule was recognized as 5.7, with relative distance (R$\sub$f/) on non-denaturing gel was 0.7. Addition of EDTA and FeCI$\sub$3/ drew the possibility that the production of hemolysin was mainly induced to overcome iron deficiency inside host animalsd infection.

• Microbiology and Biotechnology Letters
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• v.30 no.3
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• pp.230-234
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• 2002

To describe characteristics of a hemolysin in mosquitocidal Bacillus thuringiensis subsp. gyangiensis strain 21-2, Escherichia coli HB101 was transformed with a gene encoding hemolysin in the strain 21-2. Transformant 47 con-tained 2.5 kb DNA was selected by ELISA, immunoblot and DNA electrophoresis. Transformant 47-5 was recon-structed after digestion of the 2.5 kb DNA with Hind m. Transformant 47-5 contained 1.8 kb DNA and expressed 23 kDa Protein which had mosquitocidal activity to Aedes aegypti. The 23 kDa Protein itself in vitro didn't show hemolytic activity on human erythrocytes, but the protein had the activity after proteinase K treatment.

• Journal of Life Science
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• v.8 no.5
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• pp.598-603
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• 1998

A marine microbe, Vibrio anguillarum was isolated from fish and studied for its concerning pathogenic substance of hemolysin. Purification of hemolysin was achieved by the procedure of ammonium sulfate precipitation from cul-ture filtrate, DEAE-cellulose chromatography, and G-200 gel filtration with 36 fold of purification and 2.3% yield. The molecular weight of the purified hemolysin was 38,000 dalton by SDS-PAGE. The purified hemolysin was stable at pH 6-9, below 45$^{\circ}C$, and up to 1% of NaCl, respectively. $Ca^{2+}, Cu^{2+}, Zn^{2+}, Fe^{2+}$ inhibited the hemolytic activity whereas EDTA and $Mg^{2+}$ did not.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.27 no.2
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• pp.107-113
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• 1994

Among the currently recognized pathogenic vibrios, V. vulnificus and V. cholerae non O1 are the most serious bacteria from the point of view of sea food hygiene in Korea. In this paper, the authors compared the hemolytic activities of the crude hemolysin produced by V. vulnificus and V. cholerae non O1 isolated from shellfish collected in Chungmoo, Korea. The authors also attempted to improve the purification method of V. vulnificus hemolysin(VVH) and tried to make antiserum with the purified hemolysin. VVH was produced in abundance in heart infusion broth containing $2\%$ NaCl in a shaking cultivation process(140rpm) at $37^{\circ}C$ for 15 hours. While hemolysin production patterns of V. cholerae non O1 were quite different by the strain during the culture times compared with the V. vulnificus. Hemolytic activity of the VVH on sheep erythrocytes was stronger than those of rabbit, but hemolytic activities of the hemolysin produced by V. cholerae non O1 on rabbit erythrocytes were as much as twice as strong as on those of sheep and horse. VVH was purified by two steps of hydrophobic column chromatography on Phenyl-Sepharose HP with Fast Protein Liquid Chromatography(FPLC). Purification fold and yield of VVH was much improved by changing the elution buffer's pH from 6.0 to 9.8 and adding $1\%$ CHAPS(a zwitter ionic detergent) and $50\%$ ethylene glycol to the 10mM glycine buffer during the repeated hydrophobic column chromatography. Homogeneity of the purified hemolysin was shown by polyacrylamide gel electrophoresis. According to the five times repeated purification results, the specific activity was increased 27500 times and the yield was improved by $23.4\%$ on average. About $250{\mu}g$ of purified hemolysin was harvested from the 2400ml of culture supernatant of V. vulnificus. Molecular weight of VVH was estimated to be 50KDa by the SDS-PAGE and the neutralization scores of the obtained antiserum acting against VVH were $2000{\sim}8500$.

• Fisheries and Aquatic Sciences
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• v.7 no.1
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• pp.10-15
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• 2004

A total of 52 pathogenic Vibrio strains was isolated from the Gwangan Beach during summer in 2003. The isolated vibrios were composed of 6 different species: V. parahaemolyticus, V. cholerae non O1, V. fluvialis, V. vulnificus, V. alginolyticus, and V. mimicus. V. parahaemolyticus was most predominant as $46\%$ (24/52), V. cholerae non O1 was the second with $23\%$ (12/52), and V. fluvialis was the third with $17\%$ (9/52). Among the isolated strains, 22 strains showed hemolytic, proteolytic or ureolytic activity. Eight strains showed both hemolysin and protease activities, and either 6 strains showed only hemolysin activities and 7 strains only protease activities. Only one strain of V. parahaemolyticus isolates showed urease activity. The urease-positive V. parahaemolyticus strain (V. parahaemolyticus S25) showed the same biochemical characteristics as the reference strain, V. parahaemolyticus KCTC 2471 (urease­negative) except for urease production. To compare the degree of virulence of Vibrio strains having different pathogenic factors, hemolysin, protease, or urease-positive strains were injected into groups of 10 each of ICR mice (7- to l0-week-old male). The lethal rate of urease-positive V. parahaemolyticus S25 was significantly high, being $70\%$. Protease-positive strains showed $40-60\%$ of lethal rate. Hemolysin-positive strains showed no mortality, similar to non-pathogenic V. parahaemolyticus KCTC 2471 and V. parahaemolyticus FM12.

• The Korean Journal of Mycology
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• v.30 no.2
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• pp.119-123
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• 2002

In the previous studies, we surveyed 68 Korean wild mushrooms for their hemolytic activity and found that cold-water extract of Hebeloma crustuliniforme contained heat-resistant hemolysin. In this study, partially purified hemolysin of the mushroom was obtained by cold-water extraction followed by precipitation with ammonium sulfate, solubility fractionation and then dialysis. The hemolysin was found to be > 12,000 in molecular weight and its optimal hemolytic temperature was $37^{\circ}C$ and it's hemolytic activity, on washed erythrocytes and unwashed erythrocytes, respectively, was in the order of sheep > rat > human ${\geq}$ mouse > chicken and sheep > mouse > human ${\geq}$ rat > chicken. When ip injected into ICR mice at 1.38 mg/kg, it incurred prompt hemolysis as well as severe renal toxicity and hepatotoxicity. These results strongly suggest that the toxicity of Hebeloma crustuliniforme, which had been well-known as a toxic wild mushroom, may be at least partly due to its hemolysin.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.34 no.5
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• pp.502-508
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• 2001

The extracellualr hemolysin produced by pathogenic Vibrio furnissii was isolated and purified by ammonium sulfate precipitation and followed by DEAE-Sephadex A-50 (1st and 2nd) ion-exchange and Sephadex G-100 gel permeation chromatography. The molecular weight of the purified hemolysin was 61 kDa. The enzyme activity was inactivated by heating at 60 for 5 min, and inhibited by additions of $Cu^{2+},\;Zn^{2+}$ and a high concentration of cholesterol. Lysis of human erythrocytes by the enzyme was temperature dependent, and optimal temperature of the enzyme was $37^{\circ}C$. The purified hemolysin was unstable at pH 6.0, 6.5 and 10.0. The optimal concentration of human erythrocytes on hemolysins was about $1.5\%$. The purified hemolysin was active for erythrocytes from three animal species including mouse, rabbit and rat, and was the most active against rabbit erythrocytes. B blood group of human erythrocytes also showed a high sensitivity to the enzyme.

• Journal of Microbiology and Biotechnology
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• v.32 no.10
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• pp.1253-1261
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• 2022

Staphylococcus aureus (S. aureus) infection causes dramatic harm to human health as well as to livestock development. As an important virulence factor, alpha-hemolysin (hla) is critical in the process of S. aureus infection. In this report, we found that bavachin, a natural flavonoid, not only efficiently inhibited the hemolytic activity of hla, but was also capable of inhibiting it on transcriptional and translational levels. Moreover, further data revealed that bavachin had no neutralizing activity on hla, which did not affect the formation of hla heptamers and exhibited no effects on the hla thermal stability. In vitro assays showed that bavachin was able to reduce the S. aureus-induced damage of A549 cells. Thus, bavachin repressed the lethality of pneumonia infection, lung bacterial load and lung tissue inflammation in mice, providing potent protection to mice models in vivo. Our results indicated that bavachin has the potential for development as a candidate hla inhibitor against S. aureus.

• Korean Journal of Pharmacognosy
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• v.18 no.4
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• pp.216-232
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• 1987

In order to investigate the preventive and therapeutic effects of Herba Houttuyniae, experimental studies on the immune response and histological findings were undertaken. The results of these studies were summarized as follows; Delayed type hypersensitivity and rosette formation rate were significantly increased compared with control group. Hemagglutination, hemolysin titer and NK cell activity were variable, not significantly against the control group. When observed on lung tissue recovery; experimental groups were shown to be significantly recovered as compared with the control group. From these results, it is suggested that Houttuyniae Herba promotes immune response and reduces the recovery time of pneumonitis and lung abscess from the histological viewpoint.

• BMB Reports
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• v.41 no.2
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• pp.119-125
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• 2008

Our previous work confirmed that Sph2/LA1029 was a sphigomyelinase-like hemolyisn of Leptospira interrogans serogroup Icterohaemorrhagiae serovar Lai. Characteristics of both hemolytic and cytotoxic activities of Sph2 were reported in this paper. Sph2 was a heat-labile neutral hemolysin and had similar hemolytic behavior as the typical sphingomyelinase C of Staphylococcus aureus upon sheep erythrocytes. The cytotoxic activity of Sph2 was shown in mammalian cells such as BALB/C mouse lymphocytes and macrophages, as well as human L-02 liver cells. Transmission electron microscopic observation showed that the Sph2 treated BALB/C mouse lymphocytes were swollen and ruptured with membrane breakage. They also demonstrated condensed chromatin as a high-density area. Cytoskeleton changes were observed via fluorescence confocal microscope in Sph2 treated BALB/C mouse lymphocytes and macrophages, where both cytokine IL-$1{\beta}$ and IL-6 were induced. In addition, typical apoptotic morphological features were observed in Sph2 treated L-02 cells via transmission electron microscope and the percentage of apoptotic cells did increase after the Sph2 treatment detected by flow cytometry. Therefore, Sph2 was likely an apoptosis-inducing factor of human L-02 liver cells.