Hydrodynamic properties of colicin A

The hydrodynamic properties of colicin A have been studied. The molecular mass of colicin A was determined from sedimentation equilibrium centrifugation to be 63 ± 1.2 kDa, in agreement with that determined from the primary amino acid sequence [Morlon et al. (1983) J. Mol. Biol. 110 , 271–289]. The sedimentation coefficient has been analyzed over a wide range of ionic strength (NaCl 0.06 – 0.56 M) and pH (8 – 4) and was found to remain almost constant. However, below pH 5 an oligomerization of colicin A to tetramers occurred. The frictional coefficient value indicated that the shape of the colicin A monomer was very asymmetric. Analysis of the pH dependence of circular dichroism of colicin A and of its COOH‐terminal domain indicated that a sharp transition occurred between pH 4 and 3. This transition was very much reduced for the COOH‐terminal domain in the presence of a non‐ionic detergent. The presence of a lipid‐binding site in colicin A at neutral pH was demonstrated both by hydrodynamic studies with micelles of n ‐hexadecanoyl and n ‐octadecanoylphosphocholine and by differential sensitivity to a proteolytic enzyme in the presence or absence of detergent micelles. About 75 molecules of lipid were bound under these conditions suggesting that colicin A was bound to lipid micelles. In contrast, at acid pH, in the presence of an excess of lipid the tetramer was dissociated into monomers complexed to 20 – 30 lipid molecules, indicating the exposure of a high‐affinity lipid‐binding site.