Symmetrical sequences recognition

The distance between and nature of the bases in both the palindromic and the direct repeats of the recognition sequence plays an important role. It is evident that a dimeric protein would bind optimally to a twofold symmetric sequence only if the distance between the recognition elements matches the distance as determined by the protein structure. If one increases the distance by a few base pairs, a loss in cooperative binding capacity of the dimerization motifs of the protein to the rigid intervening DNA may... 

Restriction-modification is a term for bacterial enzyme systems that cleave DNA sequences. Each system consists of two distinct enzyme activities a DNA methylase and an endonuclease that catalyzes the double-strand DNA break. Type I restriction endonuclease systems have both methylase and nuclease activities in one protein molecule, which contains three subunits. One subunit contains the nuclease, one the methylase, and one a sequence recognition determinant. The recognition site is not symmetrical, and cleavage occurs some distance (up to 10 kbp) away from the recognition site, although methylation occurs within the recognition site. 

The EcoRI protein has a symmetrical globular structure with a diameter of 50 A. Each subunit is composed of a single principal domain organized according to aZ/S architecture. The domain consists of five-stranded sheets which are associated with sequence recognition and the subunit interface, six a helices including two which play a key role in DNA sequence recognition, and a major extension called the arm which wraps around the bound DNA. 

The individual domains of the two receptors both have structures similar to that of the glucocorticoid receptor, and they bind to DNA in a similar way, with their recognition helices in the major groove. The dimer contacts are, however, totally different. In the glucocorticoid receptor, which binds to a palindromic DNA sequence like the 434 repressor described in Chapter 8, the domains interact symmetrically in a head-to-head fashion equivalent... 

The consensus nucleotide sequence (see Figure 10.20b) used in the crystals is a symmetrized version of naturally occuring API recognition sites, but GCN4 binds to this sequence with a high affinity. Each half-site in this DNA is bound to one monomer of the GCN4 dimer by both sequence-specific and... 

Fig. 1.15. Bending of the DNA in the CAP protein-DNA complex. The CAP protein ( . coli) binds as a dimer to the two-fold symmetric operator sequence. The DNA is bent nearly 90deg in the complex. The turns are centered around two GT sequences (shown in black) of the recognition element.

Fig. 1.19. Tetramerization of the Lac repressor and loop formation of the DNA. The Lac repressor from E. coli binds as a dimer to the two-fold symmetric operator seqnence, whereby each of the monomers contacts a half-site of a recognition sequence. The Lac operon of E. coli possesses three operator sequences Of, 02 and 03, aU three of which are required for complete repression. Of and 03 are separated by 93 bp, and only these two sequences are displayed in the figure above. Between Of and 03 is a binding site for the CAP protein and the contact surface for the RNA polymerase. The Lac repressor acts as a tetramer. It is therefore assumed that two dimers of the repressor associate to form the active tetramer, whereby one of the two dimers is bound to 03, the other dimer binds to Of. The intervening DNA forms a so-caUed repression loop. After Lewis et al., 1996.

The HREs of the steroid hormone receptors posses a palindromic structure, comparable to the DNA binding elements of procaryotic repressors (see fig. 4.7a). The glucocorticoid receptor, for example, binds as a homodimer to the two-fold symmetrical recognition sequence, whereby the receptor is already dimerized in solution. In complex with the DNA each subimit of the dimer contacts one half-site of the HRE. As a consequence of the two-fold repeat of the recognition sequence, a high affinity binding of the receptor dimer results (compare 1.2.4). 

The recognition sequences of type II restriction endonucleases are usually symmetric they have a twofold axis of symmetry so that they read the same in the 5 -to-3 sense along opposite—but, of course, complementary—strands. Such sequences are... 

The dyad symmetry of the operator sequence is probably important in providing tight binding to two subunits of the symmetric tetrameric protein.11-13 It is also possible that repressor molecules move along DNA chains in a one-dimensional diffusion process, and that the symmetry of the operator site facilitates recognition by a protein moving from either direction.14 15... 

Listed on the left are the 16 self-complementary tetranudeotides while flanking nucleotides which expand these to the 64 self-complementary hexanucleotides head the next four columns. This arrangement is useful in looking for restriction enzymes which increase the specificity of cutting at a particular tetranucleotide. Enzymes that do not have symmetrical recognition sequences are not included. 

Restriction enzymes are endonucleases which cleave specific sequences within double-stranded DNA stretches. The recognition and cleavage sequences of a routinely used group of endonucleases (called type 11 enzymes) may be four, five, six, or eight bases long and are symmetric across the two strands of DNA. Fig. 2.12 illustrates the cleavage specificity of a widely used sixbase-motif-specific endonuclease, EcoR 1. 

Figure 9.37. Structure of the Recognition Site of ECOBN Endonuclease. (A) The sequence of the recognition site, which is symmetric around the axis of rotation designated in green. (B) The inverted repeat within the recognition sequence of AcoRV (and most other restriction endonucleases) endows the DNA site with twofold rotational symmetry.

The spectra of both wild-type lac operator 14-mer d(TGTGAGCGCTCA-CA)2 (01) as well as a number of base-pair mutants, d(Ty4TGAGCGCTCATA)2 (02) and d(TGTG7GCGCACACA)2 (03 complementary sites of mutation in the palindromic operators are italicized) have been assigned (22). These symmetrical base sequences are about two-thirds the length of the 21 base-pair wild-type sequence, and the 14-mers are believed to contain most of the important recognition sites for the lac repressor protein. 

Fig. 4.5 Oligomeric structure of nuclear receptors and structure ofthe HREs. The nuclear receptors can be subdivided into four groups based on strucutres ofthe receptors and HREs. Shown above are some representative examples, a) binding of a homodi-meric receptor to a two-fold symmetric palindromic DNA element, CR gluccocorticoid receptor, b) binding of a heterodimeric receptor to a DNA element with direct repeats ofthe recognition sequence, whereby the 5 side ofthe HRE is occupied by the...

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