Circular dichroism and molecular modeling of the E-coli TolA periplasmic domains

Colicins are killer proteins that use envelope proteins from the outer and the inner membranes to reach their cellular target in susceptible cells of Escherichia coli. Each group A colicin uses a combination of Tol proteins t o cross the outer membrane of gram-negative bacteria and to exert their kil ling activity. The TolA protein, necessary for the import of all the group A colicins, is a 421-amino acid residue protein composed of three domains ( TolAI, TolAII, and TolAIII). TolAIII interacts with the N-terminal domain o f colicin A (AT1). Analytical ultracentrifugation reveals that TolAII and T olAIII are monomer structures, TolAII has an elongated structure, and TolAI II is rather globular. Circular dichroism (CD) spectra were done with TolAI I-III, TolAII, TolAIII, AT1, and the AT1-TolAII-III complex. TolA CD spectr a reveal the presence of a-helix structure in aqueous solution and the inte nsity of the cu-helix signal is the highest with TolAII. Few structural cha nges are observed with the complex AT1-TolAII-III. Molecular modeling was d one for TolAII-III, taking into account CD and ultracentrifugation data and show that domain II can adopt a barrel structure made of three twisted a-h elices similar to coiled coil helices while domain IU: can adopt a globular structure. (C) 1999 John Wiley & Sons, Inc.