Two distinct ATP-binding domains are needed to promote protein export by Escherichia coli SecA ATPase.
Six putative ATP-binding motifs of SecA protein were altered by oligonucleotide-directed mutagenesis to try to define the ATP-binding regions of this multifunctional protein. The effects of the mutations were analysed by genetic and biochemical assays. The results show that SecA contains two essential ATP-binding domains. One domain is responsible for high-affinity ATP binding and contains motifs A0 and B0, located at amino acid residues 102-109 and 198-210, respectively. A second domain is responsible for low-affinity ATP binding and contains motifs A3 and a predicted B motif located at amino acid residues 503-511 and 631-653, respectively. The ATP-binding properties of both domains were essential for SecA-dependent translocation ATPase and in vitro protein translocation activities. The significance of these findings for the mechanism of SecA-dependent protein translocation is discussed.
Mitchell
C
C
Department of Microbiology, State University of New York at Stony Brook 11794.
Oliver
D
D
D000251:Adenosine Triphosphatases / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D000255:Adenosine Triphosphate / Q000378:metabolism*; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D001483:Base Sequence; D001665:Binding Sites; D001693:Biological Transport, Active; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D000069816:SEC Translocation Channels; D000081416:SecA Proteins; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013347:Subcellular Fractions / Q000201:enzymology
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; D000069816:SEC Translocation Channels; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D000081416:SecA Proteins