|Budget Amount *help
¥3,300,000 (Direct Cost: ¥3,300,000)
Fiscal Year 2002: ¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 2001: ¥1,400,000 (Direct Cost: ¥1,400,000)
To analyze the active region of Staphylococcus hyicus exfoliative toxin B (SHETB), mutants of His^<87> (H87A, in which His^<87> was mutated to Ala), Asp^<135> (D135A, in which Asp^<135> was mutated to Ala), Ser^<209> (S209C, in which Ser^<209> was mutated to Cys), Lys^<227> (K227E, in which Lys^<227> was mutated to Glu) and Tyr^<180> (Y180F, in which Tyr^<180> was mutated to Phe), were synthesized by site-directed mutagenesis method.
When SHETB and five mutants were inoculated to NCTC2544 cells (SHETB-susceptible cell lines) and incubated at 37℃ for 48 h, the rounding effect was appeared on the cells inoculated with SHETB, K227E and Y180F. However the rounding were not appeared on the cells inoculated with H87A, D135A and S209C. The exfoliation (Nikolsky sign) was seen in the 1-day-old chicks (SHETB-susceptible animal) inoculated with SHETB, K227E and Y180F, but not seen in the chicks inoculated with H87A, D135A and S209C.
These findings suggest that active region of SHETB is constructed by His^<87>, Asp^<135> and Ser^<209>. The regions, in which included in the above three residues, is conservative among the various serine proteases. Therefore, the toxic activity of SHETB seems to be proteolytic activity of serine protease.