The major difference between bacterial and eukaryotic supercoiling is due to the generally circular nature of bacterial chromosomes versus the linear nature of eukaryotic chromosomes and the fact that bacteria do not have nucleosomes. The cccDNA of bacteria is subject to more topological constraint and therefore tends to exist in state of greater negative supercoiling in either an interwound or spiral configuration. However, since the spiral configuration is usually associated with wrapping around a protein, this form is not as prevalent in bacteria. In bacteria, the level of supercoiling is maintained to primarily by the actions of DNA topoisomerase and DNA gyrase. In eukaryotes, negative supercoiling is achieved to a great degree via writhe in the form of left handed spirals around nucleosomes while stretches of nucleosome free DNA can engage in negative supercoiling in the interwound configuration. Nucleosomal supercoiling is controlled by a number of factors involved in chromatin remodeling including methylation and acetylation states of histones, binding of proteins to stretches of DNA altering the access to nucleosome wrapping, and interaction with the many components of the nucleosome remodeling complexes. As with bacteria, DNA topoisomerase and gyrase play a role in maintaining supercoiling in nucleosome free stretches of DNA.
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(c) Methods of Compaction The basic level of compaction in eukaryotic chromosomes is the nucleosome, a 146 nucleotide stretch of DNA wrapped around an octomer of histone proteins, with a 20-80 nucleotide linker regions in between. Chains of nucleosomal complexes are then further compacted into the 30 nm fiber in zig-zag or solenoid configuration. the 30nm fiber is then organized into 40-90kb loops held together at the base of the loop by the nuclear scaffold. Among other factors, the nuclear scaffold contains topoisomerase II (Topo II), and SMC proteins, which are chromosomal ATPases. TopoII and Smc2 and Smc4 are subunits of Condensin while Smc1, and Smc3 are part of Cohesin. Studies suggest that Condensins promote lateral compaction of chromosomes, while Cohesin promotes longitudinal compaction, through linking adjacent cohesion sites. TopoII is also a component of scaffold and colocalizes with AT-rich DNA sequences of the scaffold named SARs, which are thought to anchor DNA loops onto the chromosome axis. TopoII seems to be involved in the assembly of chromatin structure, while Condensins are required for both assembly and maintenance.
In contrast to eukarytic chomosomal organization, knowledge of bacterial chromosomal organization is much more limited. Bacteria have neither nucleosomes nor a nucleus. Rather the generally circular chromosomes are organized into compact, superhelical domains in a region called the nucleoid. The chromatin structure resembles a rosette with loops of supercoiled DNA radiating from a central core. Compaction is achieved by a combination of forces including supercoiling, compaction by proteins, transcription, and possibly RNA-DNA interactions.
2) a) Binding of proteins to DNA: Domains are like snap-on tools for proteins. They are "interchangeable" protein structures which confer specific functions on the containing proteins. In the case of DNA binding domains, they impart the protein with the ability to bind to DNA. The binding may use a variety of sequence specific and/or non-specific molecular interactions including hydrogen and ionic bonding, van de Waals forces, and hydrophobic interactions, and may involve interaction with either the major or minor grooves and/or the DNA backbone. The number of residues involved and the type and strength of bonding between the molecules varies with the particular combination of protein domain(s) and DNA sequence(s)/structure(s) to which it is bound.
DNA binding domains are generally classified into "families" which share with similar DNA binding domain properties and are grouped according to the predominant structure of the binding domain. For example: 1) HTH is 2 Î±-helices connected by a turn. The recognition helix binds in a non-sequence specific manner via hydrogen bonds and hydrophobic interactions with bases in the major groove while the other helix stabilizes the binding of the two molecules. 2) bHLH - 2 Î±-helices connected by a loop. The larger, basic helix interacts with major groove of DNA while the smaller helix functions as the dimerization domain. 3) HLH and leucine zipper motifs, an Î±-helix connected by a loop to a longer Î±-helix which may contains separate DNA binding and dimerization domains as in the leucine zipper. 4) Î²-containing - Î²-sheets, perhaps in combination with intervening loops, or forming sheets/barrells/ sandwiches, and which may use either the Î²-sheet or the loop for contact, e.g TBP and Ig-like domains. 5) Mixed Î±-/ Î²-proteins which use a mix of Î±- and Î²- structures and may contact using either or both structures, or via the intervening loops, e.g. Zinc finger proteins. It is important to note that even within a particular "family" or domain structure, the can be great variation in how the domain interacts with the DNA molecule. For example, although the Î±-helix typically inserts into the major groove parallel to the DNA backbone, many other orientations are possible and found in practice.
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In addition to sequence recognition, another function of the domain is to bring the protein and DNA into spacial proximity and achieve a conformation conducive to binding. Thus, binding often requires recognition of structural deviation such as variation from the typical B form of DNA or other structural alterations such as torsion or bending. Binding may also involve/require torsional alteration in either or both of the structures either prior to or during binding.
(b) binding of proteins to other proteins. Domains facilitate protein-protein interaction via dimerization domains, which, with the exception of the leucine zipper, are usually distinct from the DNA binding domain-add something here re:nature of dimerazation domains. Hetero- and homo- dimerization of proteins provides a method to increase the variety of target sequences, sequence specificity, and/or binding affinity. Furthermore, proteins can engage in a process called 3D domain swapping, a process by which 2 or more proteins can form a dimer or oligomer by exchanging identical structural domains. For example, the cro repressor of bacteriophage Î» uses domain-swapping to dimerize by swapping C-terminal strands.
(c) domains that activate transcription. In addition to DNA binding domains as described in a), transcription factors generally contain one or more transactivation domains, which allow them to interact with other transcription factors and/or the basal transcription machinery. Transactivation domains are generally glutamine- or proline-rich, stretches of 30-100 amino acids which enhance transcription either directly or thru recruiting of other coactivators which cannot themselves bind DNA. In addition, many transcription factors generally act as homo- or hetero-dimers and thus also contain dimerization domains.