Characterization of the finger loop domain of Ffh by random sequence libraries and sequence analysis

Abstract

Although the role of the function of the signal recognition particle (SRP) from Escherichia coli in targeting of proteins to the cytoplasmic membrane is known, details of many of its structural features require further study. The finger loop region of the Ffh protein is essential for SRP function and likely forms part of the binding site for hydrophobic membrane targeting sequences. To investigate the sequence and structural features of this domain important for its function, we have developed a genetic system to screen a random aptamer library for sequences that restored function to a finger loop deletion mutant. About 1% of random sequences can replace the wild type finger loop. Analysis of the random sequences, as well as targeted mutagenesis, shows the importance of hydrophobicity in finger loop function. All functional sequences also lack extensive secondary structural consistent with the role of the finger loop in binding a variety of ligands. Continued isolation and analysis of random sequences should provide new insights into how the finger loop functions in membrane protein targeting.