Characterization of a conserved alpha-helical, coiled-coil motif at the C-terminal domain of the ATP-dependent FtsH (HflB) protease of Escherichia coli.
FtsH (HflB) is an ATP-dependent protease found in prokaryotic cells, mitochondria and chloroplasts. Here, we have identified, in the carboxy-terminal region of FtsH (HfIB), a short alpha helix predicted of forming a coiled-coil, leucine zipper, structure. This region appears to be structurally conserved. The presence of the coiled-coil motif in the Escherichia coli FtsH (HflB) was demonstrated by circular dichroism and cross-linking experiments. Mutational analysis showed that three highly conserved leucine residues are essential for FtsH (HfIB) activity in vivo and in vitro. Purified proteins mutated in the conserved leucine residues, were found to be defective in the degradation of E. coli sigma(32) and the bacteriophage lambda CII proteins. In addition, the mutant proteins were defective in the binding of CII The mutations did not interfere with the ATPase activity of FtsH (HflB). Finally, the mutant proteins were found to be more sensitive to trypsin degradation than the wild-type enzyme suggesting that the alpha helical region is an important structural element of FtsH (HflB).
Shotland
Y
Y
Department of Molecular Genetics and Biotechnology, The Hebrew University-Hadassah Medical School, Jerusalem, P.O. Box 12272, Israel.
Teff
D
D
Koby
S
S
Kobiler
O
O
Oppenheim
A B
AB
D049069:ATP-Dependent Proteases; D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D002942:Circular Dichroism; D017124:Conserved Sequence / Q000235:genetics; D003432:Cross-Linking Reagents / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins; D006360:Heat-Shock Proteins / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009154:Mutation / Q000235:genetics; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D012808:Sigma Factor; D014157:Transcription Factors / Q000378:metabolism; D014357:Trypsin / Q000378:metabolism; D014764:Viral Proteins
D001426:Bacterial Proteins; D003432:Cross-Linking Reagents; D029968:Escherichia coli Proteins; D006360:Heat-Shock Proteins; D008565:Membrane Proteins; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D012808:Sigma Factor; D014157:Transcription Factors; D014764:Viral Proteins; C039000:cII protein, bacteriophage lambda; C069774:heat-shock sigma factor 32; D049069:ATP-Dependent Proteases; C513383:FtsH protein, E coli; D014357:Trypsin; D000251:Adenosine Triphosphatases