SECRETION AND CIRCULAR-DICHROISM ANALYSIS OF THE C-TERMINAL SIGNAL PEPTIDES OF HLYA AND LKTA

The secretion of the 107 kDa hemolysin A (HlyA) from Escherichia coli is mediated by membrane proteins hemolysin B (HlyB) and hemolysin D (H lyD). The signal for transport has been mapped to the C-terminal 60 am ino acids of the HlyA molecule. We have shown previously that the C-te rminal 70 amino acids of leukotoxin (LktA) from Pasteurella hemolytica can substitute functionally for the HlyA signal sequence. This 70 ami no acid peptide contains little primary sequence similarity to the Hly A signal sequence, and we have hypothesized that these signal sequence s assume a similar higher-order structure which is recognized by the H lyB/D transporter. In the present study, we have expressed and purifie d small peptides containing the C-terminal 61 amino acids of HlyA and the C-terminal 70 amino acids of LktA. We show that these signal pepti des are sufficient for secretion from E. coli in a HlyB/D dependent ma nner. Circular dichroism analyses show that both molecules exhibit com mon biophysical properties. In aqueous solution, they appear to be mai nly unstructured, but in a membrane mimetic environment they assume a helical secondary structure. The conformational change observed for bo th peptides going from an aqueous to a membrane mimetic environment ma y be an important feature of these signal sequences necessary for thei r recognition and transport.