Analysis of the Active Site of the A Subunit of LT Produced by Enterotoxigenic E Coli

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A plasmid encoding a mutant gene of heat-labile enterotoxin (LT), produced by enterotoxigenic Escherichia coli, - was induced by treatment of plasmid EWD 299 with hydroxylamine. A mutant strain of E. coli HB 101 carrying the mutant plasmid pTUH 6A produced a low toxic LT analogue (mutant LT), which was cross-reactive with anti-LT antibody. The mutant LT activity was less than 0.15 and 0.006% of the normal LT in the rabbit ileal loop test and in the rabbit skin peameability test, respectively. The amino acid composition of the mutant LT-B subunit was the same as that of the normal B subunit. Though the A_2 fragment of the mutant LT was identical to normal LT by DNA analysis, the A_1 fragment of the mutant LT differed from the normal A_1 fragment in one amino acid at position 112 ; namely it had lysine instead of glutamic acid from the N terminus.These data suggest that glutamic acid at position 112 from the N terminus of the A_1 fragment is important for the A subunit to express its biological activity.