Stabilizing C-terminal tails on AraC.
We examined the effects of the metabolic stability of random sequences appended to the C-terminus of the dimerization domain of the regulatory protein of the Escherichia coli arabinose operon, AraC. Genetic scoring utilized the trans dominant negative effect of the dimerization domain on the activity of intact AraC, and physical scoring used sodium dodecyl sulfate (SDS) gel electrophoresis. We confirmed previous results obtained with Arc and lambda repressors that C-terminal charged residues tend to be stabilizing and that hydrophobic residues are destabilizing. Additionally, we found that the provision of a single, charged C-terminal residue conferred significant stability that was independent of interior sequence. Hence, it appears that in the engineering of proteins, flexible tails may be freely added, with only the identity of the C-terminal amino acid being restricted. Proteins 2001;42:177-181.
Ghosh
M
M
Department of Biology, Johns Hopkins University, Baltimore, Maryland 21218, USA.
Schleif
R F
RF
D000595:Amino Acid Sequence; D051777:AraC Transcription Factor; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D011487:Protein Conformation; D012097:Repressor Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D017386:Sequence Homology, Amino Acid; D012995:Solubility; D014157:Transcription Factors
D051777:AraC Transcription Factor; C497371:AraC protein, E coli; D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D010446:Peptide Fragments; D012097:Repressor Proteins; D014157:Transcription Factors