Peptide Coupling Reagents: Carbodiimides
The other approach is to introduce backbone protecting groups which will prevent the formation of hydrogen bonds. Such protection is made by the introduction of the Hmb group on the αnitrogen . It has been shown that the presence of a Hmb unit every 6-7 residues is sufficient to disrupt the peptide aggregation . The Hmb protected amino acid is introduced under the form of N,O-bis-Fmoc-N-(2hydroxy-4-methoxybenzyl) derivative, the O-Fmoc protection being cleaved during the following piperidine treatment. At the end of the synthesis the Hmb group is cleaved in the final TFA cleavage.
Peptide Synthesizer, Automated Peptide Synthesizer
Reliable methods detecting minute amounts of unreacted amino groups are essential for the monitoring of the coupling reaction. On the other hand the ability of detecting small quantities of Fmoc peptide allows the control of the completion of the Fmoc cleavage.
One might question why one would want to prepare small proteins when one can easily express the genetic message in a variety of biological systems, including eukaryotic systems that include posttranslational modification. Limitations of normal expression systems to the 20 common amino acids restricts chemical modification of the parent protein for a variety of biophysical and pharmaceutical applications. Incorporation via molecular biology of novel amino acids as spectroscopic probes, or to introduce conformational constraints, is feasible using the suppressor mutation approach pioneered by the Schultz group (16-18). In this case, however, steric limitations of the protein biosynthetic machinery preclude the use of certain unusual amino acids or constrained dipeptides (19), which are readily accessible via organic synthesis. The quantities of protein obtained by this approach are generally small, requiring assay methods (enzymatic or spectroscopic) that are sensitive as well as specific.
Liberty Blue - Automated Microwave Peptide Synthesizer
PyAOP is the 7-aza derivative of PyBOP. As such, it is the phosphonium salt derivative of HOAt. It facilitates higher coupling yields than BOP and PyBOP. Unlike BOP, no toxic HMPA (hexamethylphosphoramide) is formed during the coupling reaction. Like PyCLOCK, PyAOP does not react with the amino terminus of the growing peptide chain und thus is useful for incorporating hindered amino acids, synthesizing difficult sequences and preparing cyclic peptides.
Nitro compound synthesis by C-C coupling - Organic …
PyCLOCK is phosphonium salt of the racemization suppressant 6-Cl-HOBt. Unlike aminium-type coupling reagents (such as HBTU, HATU), PyCLOCK does not react with free amino groups and therefore does not terminate chain elongation by forming a guanidine side product with the N-terminus of the growing peptide chain. This property renders PyCLOCK especially useful for slow couplings and for cyclization reactions. Furthermore, unlike aminium coupling reagents, PyCLOCK can be used in excess to assure a complete activation of the incoming building block’s carboxylic acid function.
Ester synthesis by esterification - Organic chemistry
The choice of one of the numerous reagents to activate the carboxyl group for nucleophilic attack by the amine function, to produce the peptide bond, depends on numerous factors. When coupling peptide fragments, reagents that minimize racemization are preferred; a traditional favorite in solution chemistry for coupling fragments has been the azide group. A common procedure is activation by a combination of a diimide, such as 1,3-diisopropylcarbodiimide (DIC) or 1,3-dicyclohexylcarbodiimide (DCCI), with hydroxybenzotriazole (HOBt), or through the use of an activated derivative of HOBt, such as TBTU, to generate the activated HOBt ester in situ. Incorporation of multiple sequential sterically hindered amino acids (valine, isoleucine, aminoisobutyric acid, etc.) often require the use of special coupling reagents, such as acid fluorides, HATU, etc.) to provide reasonable reaction rates and efficient coupling yields.
General & Introductory Chemistry
Illustrative are the solid-phase protocols for the two strategies (Boc and Fmoc) commonly used for the synthesis of peptides. The Boc strategy (Fig. 1) is often combined with a 1% to 2% cross-linked polystyrene support and a benzyl ester linkage to the polymer, requiring strong acid such as hydrogen fluoride for deprotection. The procedure favored by most synthetic laboratories uses an acid-labile linkage similar to the p-methoxybenzyl ester linkage of the Wang resin and a base-labile amino protecting group, the fluorenylmethyloxycarbonyl (Fmoc), on the added amino acids (Fig. 2). One can use side-chain protection with similar acid lability to the Wang linkage to give free peptide upon cleavage, or use more stable side-chain protection to give the protected peptide for fragment condensation after purification and characterization. In the latter case, a final deprotection with strong acid such as HF is required.