Coiled-coil leucine zipper dimerization

Ryadnov et al. (2003) have designed a coiled coil-based nanoscale linker system dubbed Belt-and-Braces. The system was novel in a number of respects. Though based on a leucine-zipper dimer design, it was a ternary system in which one peptide (the Belt ) templated the assembly of two half-sized peptides (the Braces ) thus, the system was the first and simplest example of a coiled-coil vernier assembly (Kelly et al., 1998). The Belt-and-Braces design employed all of the key design rules for a... 

Residues 50-64 of the GAL4 fragment fold into an amphipathic a helix and the dimer interface is formed by the packing of these helices into a coiled coil, like those found in fibrous proteins (Chapters 3 and 14) and also in the leucine zipper families of transcription factors to be described later. The fragment of GAL4 comprising only residues 1-65 does not dimerize in the absence of DNA, but the intact GAL4 molecule does, because in the complete molecule residues between 65 and iOO also contribute to dimer interactions. 

Figure 10.18 Side-chain interactions in the leucine zipper structure, (a) The hydrophobic side chains in spikes a and d (see Figure 10.17) form a hydrophobic core between the two coiled a helices, (b) Charged side chains in spikes and g can promote dimer formation by forming complementary charge interactions between the two a helices.

The coiled-coil structure of the leucine zipper motif is not the only way that homodimers and heterodimers of transcription factors are formed. As we saw in Chapter 3 when discussing the RNA-binding protein ROP, the formation of a four-helix bundle structure is also a way to achieve dimerization, and the helix-loop-helix (HLH) family of transcription factors dimerize in this manner. In these proteins, the helix-loop-helix region is preceded by a sequence of basic amino acids that provide the DNA-binding site (Figure 10.23), and... 

Dimerization of the Ce-zinc cluster transcription factors involves an a-helical coiled coil in the dimerization region. Coiled coils, often called leucine zippers, are also found in a large group of transcription factors that do not contain zinc. The leucine zipper is made up of two a helices in a coiled coil with every seventh residue leucine or some other large hydrophobic residue, such as isoleucine or valine. Leucine zipper transcription factors (b/zip) include factors characterized by heterodimerization, for example Fos and Jun. The a-helical DNA-binding motifs of the heterodimers recognize quite different base sequences and are continous with the a helices of the zipper. 

The leucine zipper DNA-binding proteins, described in Chapter 10, are examples of globular proteins that use coiled coils to form both homo- and heterodimers. A variety of fibrous proteins also have heptad repeats in their sequences and use coiled coils to form oligomers, mainly dimers and trimers. Among these are myosin, fibrinogen, actin cross-linking proteins such as spectrin and dystrophin as well as the intermediate filament proteins keratin, vimentin, desmin, and neurofilament proteins. 

A leucine zipper is a structural motif present in a large class of transcription factors. These dimeric proteins contain two extended alpha helices that grip the DNA molecule much like a pair of scissors at adjacent major grooves. The coiled-coil dimerization domain contains precisely spaced leucine residues which are required for the interaction of the two monomers. Some DNA-binding proteins with this general motif contain other hydrophobic amino acids in these positions hence, this structural motif is generally called a basic zipper. 

B) Helical wheel representation of residues 2-31 of the coiled coil portion of the leucine zipper (residues 249-281) of the related transcription factor GCN4 from yeast. The view is from the N terminus and the residues in the first two turns are circled. Heptad positions are labeled a-g. Leucine side chains at positions d interact with residues d and e of the second subunit which is parallel to the first. However, several residues were altered to give a coiled coil that mimics the structure of the well-known heterodimeric oncoproteins Fos and Jun (see Chapter 11). This dimer is stabilized by ion pairs which are connected by dashed lines. See John et al.172... 

Each of the monomeric proteins c-jun and c-fos, as well as other members of the leucine zipper family, has an N-terminal DNA-binding domain rich in positively charged basic amino acid side chains, an activation domain that can interact with other proteins in the initiation complex, and the leucine-rich dimerization domain.363 The parallel coiled-coil structure (Fig. 2-21) allows for formation of either homodimers or heterodimers. However, cFos alone does not bind to DNA significantly and the cjun/cFos heterodimer binds much more tightly than does cjun alone.364 The yeast transcriptional activator protein GCN4 binds to the same 5 -TGACTCA sequence as does the mammalian AP-1 and also has a leucine zipper structure.360 364 365... 

Thus, the question in coiled-coil prediction and design is what specific replacements are superimposed on the basic HPPHPPP pattern to direct the functional oligomerization state This question was first tackled by Conway and Parry, who analyzed natural coiled-coil sequences that formed dimers and trimers (Conway and Parry, 1990, 1991). Woolfson and Alber (1995) advanced this approach by comparing amino-acid profiles for these two structures directly. The work that made the biggest impact on this issue, however, was the collaborative experimental study from the Kim and Alber laboratories using the GCN4 leucine-zipper peptide model system and mutants thereof. 

Returning to the a and d sites of coiled-coil interfaces, they are not the exclusive province of hydrophobic residues polar residues are found here and, in many cases, they are highly conserved. In retrospect, this is also apparent in the amino-acid profiles of dimeric and trimeric coiled coils (Conway and Parry, 1990, 1991 Lupas et al., 1991). Furthermore, inspection of these profiles reveals trends in the data for instance, basic residues occur frequently at the a sites of dimeric coiled coils (Conway and Parry, 1990). The real importance of such inclusions, however, only became apparent with the determination of the structure of the leucine-zipper peptide GCN4-pl (O Shea et at, 1991). 

Related to these ideas, Pandya et al. (2004) have described the design of an antiparallel coiled-coil (helix-loop-helix) peptide, which is stabilized by a disulfide bridge between the termini of the peptide. Reduction of the disulfide triggered a switch to a dimeric leucine zipper. 

Moitra, J., Szilak, L., Krylov, D., and Vinson, C. (1997). Leucine is the most stabilizing aliphatic amino acid in the d position of a dimeric leucine zipper coiled coil. Biochemistry 36, 12567-12573. 

The Leucine Zipper (LZ) is a dimerization motif found in the b-LZ and b-HLH-LZ ttanscription factor families (1,2). Upon dimerization, LZs fold into parallel and two-stranded a-helical coiled-coils (3-6). The primary structure of coiled-coils forming proteins is characterized by the heptad repeats (abcdefg)n where Leu residues are conserved at positions d and positions a are mostly occupied by P-branched and hydrophobic residues while e and g positions are often occupied by acidic or basic residues (7,8). The tertiary interactions of the dimeric LZ or parallel and two-stranded a-helical coiled-coils are described by the knobs-into-holes model (3,9). 

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