Cyclization backbone

Barondeau, D. P., Kassmann, C. J., Tainer, J. A. and Getzofif, E. D. (2005). Understanding GFP chromophore biosynthesis Controlling backbone cyclization and modifying post-translational chemistry. Biochemistry 44, 1960-70. 

Stabilizing Proteins by Intein-Mediated Backbone Cyclization... 

Fig. 1.6 A IPL/EPL cyclization with the IMPACT system, B backbone cyclization using the TWIN inteins. CBD stands for chitin-binding domain. N-termini are indicated by NH2.

The disulfide bridges and, if applicable, head-to-tail backbone cyclization are indicated on top and bottom, respectively, of the generic sequence. 

The backbone cyclization and cystine knot motif together render cyclotides as a class of structurally well-defined peptides that show exceptionally high stability against chemical, thermal, or enzymatic degrada-tion. °° ° This stability has meant that they have been proposed to have applications as protein engineering templates. 

Peptide bonds are cleaved in a nonselective, but not in a completely random manner. Based on anchimeric side-chain assistance, steric factors, and bond strains, acid-labile peptide bonds are predicted to include sites containing Asp, Glu, Ser, Thr, Asn, Gin, Gly, and ProJ22l The disulfide topologies of circulin B and cyclopsychotride, backbone-cyclized peptides with three disulfide bonds, were determined by partial hydrolysis for 5 hours.[22 Occasionally, the bond between adjacent half-cystine residues is cleaved due to the nonselective nature of the mechanism of partial acid hydrolysis.[21] By this procedure, in all cases, a complex mixture of peptide fragments is produced which requires careful chromatographic separation by RP-HPLC for subsequent analysis by mass spectrometry (see Section 6.1.6.2.7). 

Iturins are a group of fungicidal cyclic lipopeptides produced by Bacillus subtilis m m m All members of this group are cyclic octapeptides with seven a-amino acids and a unique 13-amino fatty acid homologue of iturinic acid. The predominant iturin-A isomer, A2, contains the n-C14-isomer of iturinic acid (Itu) (Scheme 8). The synthesis of the molecule can be divided into two steps with the first stage consisting of the assembly of the linear octapeptide on solid phase followed by the backbone cyclization in solution. 

Due to the relatively high flexibility of cyclic peptides of larger ring size an additional cyclization is sometimes used to constrain their conformation, even by nature, e.g. amanitins and phalloidins. 316 For this purpose primarily side-chain-to-side-chain cyclization is adopted, e.g. Glu/Asp versus Lys or disulfide bridges 1322 alternatively, even the principle of backbone cyclization is applied (see Section 6.8.4)J29>80 323>3241... 

In natural bioactive peptides the modes of cyclization described previously may be prevented either by the lack of suitable side-chain functionalities for lactamization or because these as well as the amino and carboxy termini are crucially involved in the bioactivity itself, and thus cannot be modified. In order to overcome these potential limitations, the concept of backbone cyclization has been proposed.129 According to this, the cyclization is performed by a covalent interconnection of two backbone amides by artificial spacers or of one backbone amide by a correctly functionalized spacer with side-chain functions or with the N- and C-terminus of the peptide (Scheme 21). This type of strategy significantly increases the diversity of possible ring structures (see Scheme 22) and of their related libraries (see Section 6.8.4). Its potential for enhancing the stability of the related peptide derivatives toward proteolytic digestion,[417 419 potency,141942" and selectivity,11417-419 is well-established. 

Scheme 20 Building Blocks for Backbone Cyclization BU-A Protected 7V-(co-Functionalized Alkyl)amino Acids BU-B A-(co-Carboxy, co-Amino, or (o-Sulfanyl)acyl Derivatives of a Reduced Dipeptide...

Synthesis of Backbone-Cyclized Peptides General Procedure 424 ... 

For the cycloscan, conformational libraries are synthesized by cyclization of continuous or noncontinuous bioactive epitopes and not by their insertion into a scaffold. Originally, the concept of cycloscan was introduced for the generation of backbone-cyclized peptide libraries 467 however, cycloscan can also be applied to other modes of cyclization. In this approach all components of each sublibrary bear the identical sequence, and differ from each other in distinct parameters that affect their conformation, but do not alter their connectivity, and hence their potential bioactivity. This is achieved by gradually introducing discrete conformational perturbations, which allow an efficient screening of the conformational space of the parent peptide. The majority of the components of such libraries should be inactive, because they do not overlap the bioactive conformation. However, the peptide that does fit the bioactive conformation should be very potent and have all the pharmacological advantages of cyclic peptides. 

While in normal combinatorial peptide libraries (either chemical or phage display) each component has a unique sequence that is different from all others, in the cycloscan libraries all components have the same sequence, but differ in their conformation. This conformational diversity is generated in a dendrimeric hierarchy as shown exemplarily in Scheme 27 for the parent linear heptapeptide A-B-C-D-E-F-G. The diversity of the 1st order sublibrary (this nomenclature was adopted from Furka[468l) is based on the mode of cyclization. Excluding the head-to-tail cyclization there are seven different modes of cyclization that can be used for cycloscan three natural modes of cyclization and four modes of N-backbone cyclization. In addition there are five theoretical modes of C-backbone cyclization (see Scheme 1) which are not included in Scheme 27. 

Figure 16.7. Strategies for the backbone cyclization of peptides on solid phase without simultaneous...

N-alkylated a-amino acids, whereby the side chains typical of coded residues are transferred from the C - to the N-atom 4) fS-amino acids, the subclass of residues investigated most extensively in which the amino and carboxyl functionalities are separated by more than one carbon atom and 5) short-range, backbone-to-backbone cyclizations that generate small-ring, monolactam or dilactam building blocks. 

Cyclization is one of the earliest techniques applied to design peptidomimetics. Cyclic peptides are more stable to amide bond hydrolysis and allow less conformational flexibility consequently, the resulting analogs are anticipated to be more selective and less toxic. Methods for restricting conformations include peptide backbone cyclization, disulfide bond formation, side-chain cyclization, and metal ion chelation. 

The two most common reactions for the generation of cyclic peptides are (i) disulfide formation between cysteine or penicillamine residues through oxidation, and (ii) lactam formation between the amino and carboxy terminal ends (head-to-taU cyclization), the annino terminus and the side chain of an aspartic or glutanoic add residue, or between the side chains of a lysine (or another diamino acid) and aspartic or glutanoic acid residues. A special mode of the second type is backbone cyclization between an amino group (N-terminal or side chain) and a carboxyalkylated backbone nitrogen. Both reactions have been used to generate cyclic peptide libraries. 

Cyclization reaction, approach to the synthesis of cyclic peptides. Cyclization reactions comprise backbone cyclization (head-to-taU cyclization), side chain-to-side chain ring closure, side chain-to-head and... 

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