A thermodynamic model of the cooperative interaction between the archaeal transcription factor Ss-LrpB and its tripartite operator DNA

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• 作者：Eveline Peeters^a ; ^{espeeter@vub.ac.be} ; Liesbeth van Oeffelen^a ; ^b ; Marc Nadal^c ; ^d ; Patrick Forterre^c ; ^e ; Daniel Charlier^a
• 关键词：Bp ; base pair ; BRE ; factor B recognition element ; Cu-OP ; copper-phenantroline ; EMSA ; electrophoretic mobility shift assay ; FL ; full-length ; HTH ; helix-turn-helix ; KBi ; equilibrium association constant for binding at box Bi ; Ki ; stepwise equilibrium consta
• 刊名：Gene
• 出版年：2013
• 出版时间：25 July, 2013
• 年：2013
• 卷：524
• 期：2
• 页码：330-340
• 全文大小：1375 K

文摘

Ss-LrpB is a transcription factor of the archaeon Sulfolobus solfataricus that belongs to the leucine-responsive regulatory protein family. This protein binds to three distinct binding sites in the control region of its own gene, suggestive of autoregulation. Here, we present a detailed study of the thermodynamic and conformational rules that govern the interaction between Ss-LrpB and its tripartite operator DNA. Lane-per-lane partition analysis of macroscopic binding state populations in electrophoretic mobility shift assays, probing binding to full-length, truncated and mutated forms of the operator, allowed determination of equilibrium association constants and cooperativity parameters. The resulting thermodynamic model demonstrates that the Ss-LrpB-operator regulatory complex is formed with a significant positive cooperativity, which is mostly arising from dimer-dimer interactions between pairs of adjacent binding sites. There is a constraint on the spacing between these binding sites, with a preference for a cis-alignment on the DNA helix and with a 16-bp linker yielding maximal pairwise cooperativity. DNase I footprinting assays demonstrated that the extent of Ss-LrpB-induced DNA deformations depends on linker length. The knowledge of the thermodynamic principles underlying the Ss-LrpB-operator interaction, presented here, will contribute to unraveling of the cis-regulatory code of Ss-LrpB autoregulation.