FC0052
High-mobility protein group B1 (HMGB1)  -  DNA

Biological function
High mobility group (HMG) protein B1 (HMGB1) participates in various nuclear processes. HMGB1 bends DNA significantly, and it appears to act primarily as an architectural facilitator in the assembly of nucleoprotein complexes, in which the bound DNA is often tightly bent. Histone H1 and the non-histone high-mobility group protein B1 (HMGB1) are chromatin components that have opposite effects on chromatin stability. H1, present at one molecule per nucleosome on average, stabilizes both the nucleosome and chromatin higher-order structure, whereas HMGB1, which is about 10-fold less abundant, is a potential destabilizer and may act in conjunction with chromatin remodelling complexes. Both HMGB1 and H1 are highly mobile in vivo, with high on/off rates of association with chromatin. This mobility could provide windows of opportunity during which one protein could replace the other in chromatin. For example, HMGB1 could effectively displace H1 by competing for its binding site, or part of it, resulting in local destabilization of chromatin and possibly recruitment of other proteins, leading to transcriptional activation.

Domain organization/sequence features
HMGB1 has a tripartite structure consisting of two homologous tandem HMG-box DNA-binding domains of ∼80 amino-acid residues (the A and B boxes) linked by a 20-residue basic extension to a 30-residue C-terminal acidic tail composed entirely of aspartic acid and glutamic acid residues. The HMG box binds to linear DNA in the minor groove with little or no sequence specificity, inducing a significant bend in the DNA, and has a preference for distorted DNA structures. HMGB1 binding to target DNA via the two HMG boxes is negatively regulated by the disordered C-terminal tail.

Structural evidence
By CD measurements the tail in full-length HMGB1 adopts a random-coil conformation, and the secondary and tertiary structure are unaffected by tail length in the truncated HMGB1 variants. NMR experiments show that a large proportion of the tail resonances shift, indicating that the tail makes extensive contacts with the body of the protein. The proximal half of the tail (residues 185–199) primarily contacts the N- and C-terminal regions of each box, and the concave face of the B box. The next ten residues (200–209) contact the concave face of the A box. The five residues that extend the tail from Δ15 to Δ10 (residues 200–204) appear to be involved directly in recruiting the A box to the assembly. Long- range NOEs between the tail and the boxes/linkers indicate that it does not appear to adopt any stable secondary structure. The tail-bound states must be a dynamic equilibrium between the more compact, tail-bound form(s) and ’open’, free forms in which the DNA-binding surfaces are accessible. Relaxation enhancements indicate the existence of a ’sandwich’-like collapsed structure in which the tail enables the close approach of the basic domains.

Biochemical evidence
A series of HMGB1 tail-truncation mutants differing from each other by five residues was generated. Chemical-shift perturbation mapping using these mutants shows that tails of different lengths bind with different affinities. The acidic tail also stabilizes the protein against thermal denaturation.

Structure/Mechanism
The disordered C-terminal tail stabilizes HMGB1, but does not perturb its secondary or tertiary structure. In the absence of DNA, the two HMG boxes assemble on the acidic C-terminal tail and the binding surfaces are only transiently exposed. The tail-bound collapsed form is in dynamic equilibrium with an extended DNA binding competent form, in which the tail remains disordered. The tail screens interactions between the two HMG boxes, therefore, it affects DNA recognition in a length- dependent manner. The tail negatively regulate the affinity of the HMG boxes for most DNA substrates. When the tail is displaced by DNA it would become available for possible interaction with histones or other proteins. In addition, proteins interacting with the acidic tail in free HMGB1 may liberate the boxes and enhance their binding to DNA. It is worth noting that, despite its low level of sequence complexity and uniformly acidic character, the tail binds in a specific, but nonetheless largely or wholly electrostatic manner.

Mechanism category
competitive binding

Significance
Dynamic association of HMGB1 with chromatin and provide a mechanism by which protein–protein interactions or posttranslational modifications might regulate the function of the protein at particular sites, or at particular stages in the cell cycle.