Conformational changes of a Swi2/Snf2 ATPase during its mechano-chemical cycle

Abstract

Remodelling protein nucleic acid interfaces is an important biological task, which is often carried out by nucleic acid stimulated ATPases of the Swi2/Snf2 superfamily. Here we study the mechano-chemical cycle of such an ATPase, namely the catalytic domain of the Sulfolobus solfataricus Rad54 homologue (SsoRad54cd), by means of fluorescence resonance energy transfer (FRET). The results of the FRET studies show that the enzyme can be found in (at least) two different possible conformations in solution. An open conformation, consistent with a recently reported crystal structure, is converted into a closed conformation after DNA binding. Upon subsequent binding of ATP no further change in conformation can be detected by the FRET measurements. Instead, a FRET detectable conformational change occurs after ATP hydrolysis and prior to ADP release, suggesting a powerstroke that is linked to phosphate release. Based on these data we will present a new model for the mechano-chemical cycle of this enzyme. This scheme in turn provides a working model for understanding the function of other members of the Swi2/Snf2 family.

Two-step Mechanism for Modifier of Transcription 1 (Mot1) Enzyme-catalyzed Displacement of TATA-binding Protein (TBP) from DNA

“…5A) implicates direct Mot1-DNA interactions in contributing to the stability of the Mot1-TBP-DNA ternary complex in the absence of nucleotide. The measured affinity for the Mot1C-DNA interaction described here is similar to that observed for the ATPase domain of another Snf2/Swi2 family member, SsoRad54 (41). Together, sequence conservation, DNase I footprinting experiments (43), structural studies of the Mot1-TBP complex (35) and the direct observation of DNA binding activity of this study for Mot1C provide strong support for positioning the ATPase domain of Mot1 on DNA upstream of the TATA box.…”

“…Upon ATP binding (step 3), the ATPase domain of Mot1 is proposed to undergo a conformational change (41), resulting in a decrease in affinity for DNA (Fig. 5C).…”

“…Mot1-TBP complexes have relaxed sequence specificity for DNA (36), but the cause of this altered behavior is unclear. In contrast to other Snf2/Swi2 family members (37)(38)(39)(40)(41), full-length Mot1 has no detectable DNA binding affinity (16,36,42,43), and the ATPase activity of Mot1 is stimulated by TBP in vitro (44,45). However, biochemical evidence indicates that the ATPase domain is in close proximity to DNA when Mot1 is assembled with TBP-DNA (35,43), suggesting direct Mot1-DNA interaction.…”

Regulation of the Rhp26 ERCC6/CSB chromatin remodeler by a novel conserved leucine latch motif

“…The opening and closing of the interlobe cleft and relative rotation between lobe 1 and lobe 2 during the ATP hydrolysis cycle is proposed to cause the relative DNA-affinity change of the two lobes, resulting in the translocation along the DNA and nucleosome (chromatin remodeling) (23,24,30). Our chemical cross-linking data suggest that the leucine latch motif is likely to dock at a specific area at lobe 2/HD2 of Rhp26.…”

Molecular Mechanism of Mot1, a TATA-binding Protein (TBP)-DNA Dissociating Enzyme

“…Crystal structures of SF2 ATPases, as well as related SF1 superfamily members, have revealed the presence of two types of conformational states, "open" and "closed," whose interconversion appears to be mediated by the binding and hydrolysis of ATP (26, 28 -30). Biochemical evidence for nucleotide-mediated conformational changes inferred from the structural data was obtained in fluorescence resonance energy transfer (FRET) studies using the Snf2/Swi2-related enzyme SsoRad54 (31). Importantly, emerging evidence provides support for the idea that the conversion between such conformational states is important for ATPase regulation (27).…”