tailieunhanh - Báo cáo Y học: DNA supercoiling in Escherichia coli is under tight and subtle homeostatic control, involving gene-expression and metabolic regulation of both topoisomerase I and DNA gyrase

DNA of prokaryotes is in a nonequilibrium structural state, characterized as ÔactiveÕ DNA supercoiling. Alterations in this state a ect many life processes and a homeostatic control of DNA supercoiling has been suggested [Menzel, R. & Gellert, M. (1983) Cell 34, 105±113]. We here report on a new method for quantifying homeostatic control of the high-energy state of in vivo DNA. The method involves making small perturbation in the expression of topoisomerase I, and measuring the e ect on DNA supercoiling of a reporter plasmid and on the expression of DNA gyrase | Eur. J. Biochem. 269 1662-1669 2002 FEBS 2002 DNA supercoiling in Escherichia coh is under tight and subtle homeostatic control involving gene-expression and metabolic regulation of both topoisomerase I and DNA gyrase Jacky L. Snoep1 2 Coen C. van der Weijden1 Heidi W. Andersen2 3 Hans V. Westerhoff1 4 and Peter Ruhdal Jensen3 1Departments of Molecular Cell Physiology and Mathematical Biochemistry BioCentrum Amsterdam Free University Amsterdam the Netherlands department of Biochemistry University of Stellenbosch South Africa 3Section of Molecular Microbiology Biocentrum Technical University of Denmark Lyngby Denmark 4Stellenbosch Institute for Advanced Study South Africa DNA of prokaryotes is in a nonequilibrium structural state characterized as active DNA supercoiling. Alterations in this state affect many life processes and a homeostatic control of DNA supercoiling has been suggested Menzel R. Gellert M. 1983 Cell 34 105-113 . We here report on a new method for quantifying homeostatic control of the high-energy state of in vivo DNA. The method involves making small perturbation in the expression of topoisomerase I and measuring the effect on DNA supercoiling of a reporter plasmid and on the expression of DNA gyrase. In a separate set of experiments the expression of DNA gyrase was manipulated and the control on DNA supercoiling and topoisomerase I expression was measured part of these latter experiments has been published in Jensen . van der Weijden . Jensen . Westerhoff . Snoep . 1999 Eur. J. Biochem. 266 865-877 . Of the tWo regulatory mechanisms via which homeostasis is conferred regulation of enzyme activity or regulation of enzyme expression we quantified the first to be responsible for 72 and the latter for 28 . The gene expression regulation could be dissected to DNA gyrase 21 and to topoisomerase I 7 . On a scale from 0 no homeostatic control to 1 full homeostatic control we quantified the homeostatic control of DNA supercoiling at . A

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