TY - JOUR
T1 - Functional dissection of a cell-division inhibitor, SulA, of Escherichia coli and its negative regulation by Lon
AU - Higashitani, A.
AU - Ishii, Y.
AU - Kato, Y.
AU - Horiuchi, K.
N1 - Funding Information:
Acknowledgements We thank Nahoko Higashitani and Emiko Suzuki for their help in experiments, and Hiroshi Hara for critical reading of the manuscript. This work was supported in part by grants-in-aid from the Ministry of Education, Science and Culture of Japan.
PY - 1997
Y1 - 1997
N2 - SulA is induced in Escherichia coli by the SOS response and inhibits cell division through interaction with FtsZ. To determine which region of SulA is essential for the inhibition of cell division, we constructed a series of N-terminal and C-terminal deletions of SulA and a series of alanine substitution mutants. Arginine at position 62, leucine at 67, tryptophan at 77 and lysine at 87, in the central region of SulA, were all essential for the inhibitory activity. Residues 3-27 and the C-terminal 21 residues were dispensable for the activity. The mutant protein lacking N-terminal residues 3-47 was inactive, as was that lacking the C-terminal 34 residues. C-terminal deletions of 8 and 21 residues increased the growth-inhibiting activity in lon+ cells, but not in lon- cells. The wild-type and mutant SulA proteins were isolated in a form fused to E. coli maltose-binding protein, and tested in vitro for sensitivity to Lon protease. Lon degraded wild-type SulA and a deletion mutant lacking the N-terminal 93 amino acids, but did not degrade the derivative lacking 21 residues at the C-terminus. Futhermore, the wild-type SulA and the N-terminal deletion mutant formed a stable complex with Lon, while the C-terminal deletion did not. MBP fused to the C-terminal 20 residues of SulA formed a stable complex with, but was not degraded by Lon. When LacZ protein was fused at its C-terminus to 8 or 20 amino acid residues from the C-terminal region of SulA the protein was stable in lon+ cells. These results indicate that the C-terminal 20 residues of SulA permit recognition by, and complex formation with, Lon, and are necessary, but not sufficient, for degradation by Lon.
AB - SulA is induced in Escherichia coli by the SOS response and inhibits cell division through interaction with FtsZ. To determine which region of SulA is essential for the inhibition of cell division, we constructed a series of N-terminal and C-terminal deletions of SulA and a series of alanine substitution mutants. Arginine at position 62, leucine at 67, tryptophan at 77 and lysine at 87, in the central region of SulA, were all essential for the inhibitory activity. Residues 3-27 and the C-terminal 21 residues were dispensable for the activity. The mutant protein lacking N-terminal residues 3-47 was inactive, as was that lacking the C-terminal 34 residues. C-terminal deletions of 8 and 21 residues increased the growth-inhibiting activity in lon+ cells, but not in lon- cells. The wild-type and mutant SulA proteins were isolated in a form fused to E. coli maltose-binding protein, and tested in vitro for sensitivity to Lon protease. Lon degraded wild-type SulA and a deletion mutant lacking the N-terminal 93 amino acids, but did not degrade the derivative lacking 21 residues at the C-terminus. Futhermore, the wild-type SulA and the N-terminal deletion mutant formed a stable complex with Lon, while the C-terminal deletion did not. MBP fused to the C-terminal 20 residues of SulA formed a stable complex with, but was not degraded by Lon. When LacZ protein was fused at its C-terminus to 8 or 20 amino acid residues from the C-terminal region of SulA the protein was stable in lon+ cells. These results indicate that the C-terminal 20 residues of SulA permit recognition by, and complex formation with, Lon, and are necessary, but not sufficient, for degradation by Lon.
KW - Cell cycle control
KW - FtsZ
KW - Protease
KW - Protein recognition
KW - SOS response
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U2 - 10.1007/s004380050426
DO - 10.1007/s004380050426
M3 - Article
C2 - 9180687
AN - SCOPUS:0030943075
VL - 254
SP - 351
EP - 357
JO - Zeitschrift für Induktive Abstammungs- und Vererbungslehre
JF - Zeitschrift für Induktive Abstammungs- und Vererbungslehre
SN - 1617-4615
IS - 4
ER -