Two precursors of the heat‐stable enterotoxin of Escherichia coli: evidence of extracellular processing

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Abstract
Expression of the gene of the methanol‐soluble, heat‐stable enterotoxin of Escherichia coli (STA) allowed the identification by SDS‐PAGE of a cell‐associated 7500 Dalton STA‐related peptide; when similar experiments were performed with a phosphate buffer SDS‐PAGE system, an additional Mr9800 band became apparent. The 9800 Dalton form, pre‐pro‐STA, accumulated as an intracellular species when the experiments were performed in the presence of the proton ionophore CCCP (carbonylcyanide m‐chlorophenylhydrazone); by pulse‐chase experiments, it was shown that pre‐pro‐STA became a periplasmic Mr 7500 pro‐STA and this form was chased to the culture supernatant; periplasmic and extracellular pro‐STA showed the same electrophoretic mobility. A short time after the pulse, pro‐STA was converted extracellularly to mature STA (Mr 4500). It is proposed that STA is synthesized as pre‐pro‐STA, a 72‐amino‐acid peptide that is subsequently cleaved between amino acids 19 and 20as it is translocated across the inner membrane. The resulting 53‐amino‐acid pro‐STA is first detected in the periplasm and is then secreted to the culture supernatant. Pro‐STA is cleaved extracellularly to yield mature STA (Mr 4500).