P-Tum formation

In the heavy-chain hinge fragment 225-232 the intervening sequence between the two cysteine residues is Pro-Pro and the native structure of this portion of the IgGl molecule consists of a parallel dimer. By estimating for the Cys-Pro-Pro-Cys portion the propensity for chain reversal, l.e. for p-tum formation, according to... 

Uny et also reported the chemical synthesis of protein polymers based on the (Val-Pro- Ala-Val-Gly) repeat sequence in which glycine is replaced by the D-alanine residue. The hetero-chiral Pro- Ala diad would be erqrected on the basis of stereochemical considerations to adopt a type-II p-tum conformation. Stmctural analyses of small-molecule "Pro- Ala turn models support the formation of the type-II p-mm conformation in solution and the solid state. Polymers based on the (Val-Pro- Ala-Val-Gly) repeat sequence display a thermo-reversible phase transition similar to the corresponding polypeptides derived from the parent (Val-Pro-Gly-Val-Gly) sequence, albeit with a shift of the Tt to approximately 5-10 ° G below the latter due to a slight inaease in hydrophobic character due to the presence of the alanine residue. NMR spectroscopic analyses of the (Val-Pro- Ala-Val-Gly) polymer suggest that the repeat unit retains the p-tum stmcture on the basis of comparison to the corresponding behavior of the (Val-Pro-Gly-Val-Gly) polymer. Stress-strain measurements on cross-linked matrices of the (Val-Pro- Ala-Val-Gly) polymer indicate an elastomeric mechanical response in which the elastic modulus does value in comparison to the (Val-Pro-Gly-Val-Gly) polymer. These smdies of glycine suhstitution support the hypothesis that type-II p-tum formation can he associated with the development of elastomeric behavior with native elastins and elastin-derived polypeptide sequences. Several investigators have proposed that the (Val-Pro-Gly-Val-Gly) pentapeptide represents the minimal viscoelastic unit... 

Golebiowski A, Jozwik J, Klopfenstein SR, Colson AO, Grieb AL, Russell AF, Rastogi VL, Diven CP, Portlock DE, Chen JJ (2002) Solid-supported synthesis of putative peptide P-tum mimetics via Ugi reaction for diketopiperazine formation. J Comb Chem 4 584—590... 

Scheme 47 Formation of a Lactam p-Tum Peptidomimetic and Its Use in Preparation of the Substance P Mimetid164 ...

Solid-state 13C NMR has been shown to be a more effective analytical tool for demonstrating the formation of P-sheets in polypeptides and proteins, because the isotropic 13C NMR chemical shifts of carbon atoms in proteins are sensitive to the P-sheet s secondary structure. It is well established that SF conformations are dependent upon the species of silkworms and conditions of the sample preparation. In particular, has been reported that fibroin from Bomhyx mori adopts two dimorphic structures, silk I and silk II. The silk II form is identified by the C chemical shifts of glycine (Gly), serine (Ser), and alanine (Ala) that are indicative of P-sheets, while the silk I form produces chemical shifts that are associated with a loose helix or distorted P-tum. However, when compared with silk II, the less stable silk I shows a relatively unresolved structure, and the conformation of the soluble form of SF rapidly undergoes a transition to the insoluble silk II conformation. 

The tenet of classical rubber theory has been that the chains are in random networks and the networks comprise a Gaussian distribution of end-to-end chain lengths. However, the mechanisms and molecular bases for the elasticity of proteins are more complex than that of natural rubber. In biological systems elastomeric proteins consist of domains with blocks of repeated sequences that imply the formation of regular stmctures and domains where covalent or noncovalent cross-linking occurs. Although characterised elastomeric proteins differ considerably in their precise amino acid sequences they all contain elastomeric domains comprised of repeated sequences. It has also been suggested that several of these proteins contain p-tums as a structural motif (Tatham and Shewry 2000). 

Peptide models of a-helices have revealed much about the intrinsic and extrinsic factors that control helix formation in peptides and proteins (1-5). While considerable progress has been made in our understanding of helix formation and stabilization, the same cannot be said of the situation regarding two other important classes of secondary structure p-sheets and p-tums. The reason for this is that there has not yet been a p-sheet model developed that is as simple to prepare and as easy to characterize as a monomeric helix. 

The formation of a fourth cytoplasmic loop by palmitoylation of cysteine residues in the C-terminus has been shown to be crucial in G-protein coupling for some receptor subtypes. Yeagle et al constructed a polypeptide containing 43 amino acid residues of the carboxyl terminal domain of bovine rhodopsin and performed a similar study as previously described. The representative stmeture of the carboxyl terminal shows an organisation of the stmeture into three subdomains. The first subdomain, the N-terminal domain of this peptide, forms an a-helix containing 7 or 8 amino acids, which is believed to be a continuation of helix 7. The second subdomain contains the two cysteine residues, which are situated near the location of a putative lipid bilayer and therefore are accessible to palmitoylation. With the location of the cysteine residues near to the putative lipid bilayer, a loop is formed between the cysteine residues and helix 7. The third subdomain is partially composed of an antiparallel P-sheet with the two strands connected by a p-tum. This subdomain appeared to be the most exposed, which makes the phosphorylation sites readily available to rhodopsin kinase . 

The most obvious structural building blocks in proteins are the elements of secondary structure, the a-helix and the P-strand. These elements arise, in part, through the formation of intramolecular hydrogen bonds. Secondary structure prediction usually attempts to assign the conformational state of a residue as helix (H), extended (E), or coil (C), based on the local sequence (i.e., the surrounding residues). However, other ordered structures, such as P-tums and Q-loops, do exist. Secondary... 

The hairpin-like P-turn has recently been examined in more detail from the standpoint of conformational energy (28). This investigation revealed that there are two types of conformation of the L L bend which accomodate the sequences L -> L, L G, G - L, G G, while only one type is possible for the L D bend which accomodates the sequences L - D, L G, G D, G G. The new investigation supported and extended the earlier considerations, and has been successfully compared with the experimental data. These investigations revealed that the formation of the /1-turn is a determining factor in the cyclization of linear peptides and explained why a particular amino acid such as D-amino acid or a constrained amino acid like proline are often found at the comer of the p-tum in thetic and naturally occurring cyclic peptides. 

Supported reagents were further applied to achieve post-cleavage purification of products obtained by solid-phase synthesis. This is illustrated by an example from Ellman s laboratory Scheme 1.6.22). The p-tum mimetic precursor 43 was released from solid support by TCEP (41) mediated cleavage of a disulfid linkage. The polymeric guanidine 44 was used to remove excess of 41 and of the phosphinoxide 42, to promote formation of the cyclic sulfide 45, and to trap the byproduct HBr from solution. 

Figure 8 Ball and stick diagram of the Val-Pro-Gly-Val sequence that defines a type-ll p-tum within the crystallographically determined structure of cyclo(Val-Pro-Gly-Val-Gly)3 (CCDC ID VPGVGB10). Note that the Vaf (CO) (HN)Val distance of 3.28 A is long compared to that typically observed for hydrogen-bonded structures within proteins (-2.9 A). In contrast, the Val 0) Pro (C) distance is relatively short, 2.77 A, and the nonbonded angle O (C=0) is relatively obtuse, 99.4°. These structural features are consistent with the presence of an n n interaction that stabilizes the type-ll p-tum within these elastin repeat sequences in combination with the n a interaction of hydrogen bond formation. Note the Cy-exo conformation of the Pro residue of the type-ll p-turn sequence.

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