TY - JOUR
T1 - Diversity of the metalloprotease toxin produced by enterotoxigenic Bacteroides fragilis
AU - Wu, Shaoguang
AU - Dreyfus, Lawrence A.
AU - Tzianabos, Art O.
AU - Hayashi, Chika
AU - Sears, Cynthia L.
PY - 2002
Y1 - 2002
N2 - Enterotoxigenic Bacteroides fragilis (ETBF) strains produce a 20-kDa zinc metalloprotease toxin (BFT) associated with diarrheal disease of animals, young children, and adults. BFT stimulates secretion in intestinal loops in vivo and modifies epithelial cell morphology in vitro. The B. fragilis toxin (bft) gene from ETBF strain 86-5443-2-2 (piglet; bft-2) revealed significant nucleotide and predicted amino acid differences when compared to the bft gene from ETBF strain VPI 13784 (lamb; bft-1). This study compares BFT-1 and BFT-2, respectively, produced by ETBF strains VPI 13784 and 86-5443-2-2 purified using the Van Tassell method (38) and a modified purification scheme described herein. Multiple differences in the protein toxins produced by these ETBF strains were identified. First, purified BFT-1 eluted from a high-resolution anion-exchange column (Mono Q) at 0.22 ± 0.005 M NaC1 versus 0.18 ± 0.001 M NaC1 for BFT-2 (P < 0.001). Second, BFT-1 and BFT-2 exhibited different electrophoretic mobilities on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and reverse-phase fast protein liquid chromatography. Third, each BFT reacted with greater specificity to homologous rather than heterologous antisera. Fourth, BFT-2 had modest, but consistently, greater biological activity than BFT-1 when tested on HT29/C1 cells (P ≤ 0.01). Together, these data indicate that these ETBF strains produce two distinct isotypes of BFT, termed BFT-1 (VPI 13784 BFT) and BFT-2 (86-5443-2-2 BFT) to recognize the order in which the proteins were purified and genetic sequences identified. The modified purification scheme described in this report yields about two to three times more purified BFT protein than previous protocols and is less time consuming.
AB - Enterotoxigenic Bacteroides fragilis (ETBF) strains produce a 20-kDa zinc metalloprotease toxin (BFT) associated with diarrheal disease of animals, young children, and adults. BFT stimulates secretion in intestinal loops in vivo and modifies epithelial cell morphology in vitro. The B. fragilis toxin (bft) gene from ETBF strain 86-5443-2-2 (piglet; bft-2) revealed significant nucleotide and predicted amino acid differences when compared to the bft gene from ETBF strain VPI 13784 (lamb; bft-1). This study compares BFT-1 and BFT-2, respectively, produced by ETBF strains VPI 13784 and 86-5443-2-2 purified using the Van Tassell method (38) and a modified purification scheme described herein. Multiple differences in the protein toxins produced by these ETBF strains were identified. First, purified BFT-1 eluted from a high-resolution anion-exchange column (Mono Q) at 0.22 ± 0.005 M NaC1 versus 0.18 ± 0.001 M NaC1 for BFT-2 (P < 0.001). Second, BFT-1 and BFT-2 exhibited different electrophoretic mobilities on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and reverse-phase fast protein liquid chromatography. Third, each BFT reacted with greater specificity to homologous rather than heterologous antisera. Fourth, BFT-2 had modest, but consistently, greater biological activity than BFT-1 when tested on HT29/C1 cells (P ≤ 0.01). Together, these data indicate that these ETBF strains produce two distinct isotypes of BFT, termed BFT-1 (VPI 13784 BFT) and BFT-2 (86-5443-2-2 BFT) to recognize the order in which the proteins were purified and genetic sequences identified. The modified purification scheme described in this report yields about two to three times more purified BFT protein than previous protocols and is less time consuming.
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U2 - 10.1128/IAI.70.5.2463-2471.2002
DO - 10.1128/IAI.70.5.2463-2471.2002
M3 - Article
C2 - 11953383
AN - SCOPUS:0036120339
VL - 70
SP - 2463
EP - 2471
JO - Infection and Immunity
JF - Infection and Immunity
SN - 0019-9567
IS - 5
ER -