Methods of Cleavage

Partial acid hydrolysis is generally non-specific but can still be exploited for the cleavage of the peptide bond at aspartyl residues. The Asp-Pro linkage is exceptionally sensitive to acids and is cleaved by dilute hydrochloric acid at room temperature. At elevated temperatures very weak acids such as 0.03 M 

HCl or 0.25 M acetic acid are sufficient to effect hydrolysis of the peptide bonds on both sides of aspartyl residues. For instance the hexapeptide Val-Leu-Gly-Asp-Phe-Pro yields the tripeptide Val-Leu-Gly, the dipeptide Phe-Pro and free aspartic acid. Release of aspartic acid is usually complete after a day of hydrolysis at 100 °C in 0.25 M AcOH while hydrolytic fission of other peptide bonds is negligible. Asparagine creates a notable exception because the carboxamide group in its side chain is fairly sensitive to acid catalyzed hydrolysis and the free carboxyl group of the gradually formed aspartyl residue provides the neighboring group effect which leads to the excision of aspartic acid from the chain. 

The reaction is readily monitored by the characteristic changes in the uv spectrum. A more reliable source of positively charged bromine is N-bromosuccin-imide, which has often been used for cleavage at tryptophan  

Of course the tyrosine side-chain is not inert toward N-bromosuccinimide and to some extent also histidine residues are affected. A more selective reagent for the cleavage at tryptophan was sought and found in the skatol derivative 

A somewhat analogous fission of the peptide chain occurs when methionine containing peptides are exposed to the action of cyanogen bromide  

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For other methods of cleavage the chapter on alcohol protection should be consulted. 

Later, the method of cleavage of the tertiary acetylenic alcohols in boiling benzene or toluene in the presence of NaOH or KOH was reported (88M253 2001UP1). 

Selective cleavage of peptides and proteins is an important procedure in biochemistry and molecular biology. The half-life for the uncatalyzed hydrolysis of amide bonds is 350 500 years at room temperature and pH 4 8. Clearly, efficient methods of cleavage are needed. Despite their great catalytic power and selectivity to sequence, proteinases have some disadvantages. Peptides 420,423,424,426 an(j proteins428 429 can be hydrolytically cleaved near histidine and methionine residues with several palladium(II) aqua complexes, often with catalytic turnover. 

Most of the reagents required for the saponification of resin-bound esters are, unfortunately, non-volatile, and purification of the products is generally necessary. This is probably the main reason for the scarcity of examples of cleavage by saponification as compared with other methods of cleavage. 

Trichloroethylene (TCE) and perchloroethylene (PCE) require cleavage of the carbon-halogen bonds. Two methods of cleavage are P-elimination by dehy-drohalogenation, as shown in Equation (13.14), and nucleophilic substitution by either water or hydrogenolysis in Equation (13.15). The proposed pathways for reduction of chloroethylenes by zero-valent iron are as follows ... 

CF3C02H, CH2C12, 23°, 1 h, 74% yield.1 ( 2. For other methods of cleavage the chapter on alcohol protection should be consulted. 

A more direct method of cleavage has been developed which consists of heating 502 overnight in toluene with a dispersion of sodium and chlorotrimethylsi-lane.352 418 This procedure led efficiently to 506 whose hydrolysis with dry methanol (kinetic quenching) resulted in the production of a mixture of dihydro diesters rich in 507 (77 %). The structurally rigid 508 and related a-diketones have subsequently been elaborated from this diester as starting material.418) Related endo.endo diesters bearing external methyl and methylthio substituents (509) have also been synthesized.419 ... 

A promising SP method of cleavage that is becoming more common is the so-called cyclative cleavage (CC). SPS produces an advanced open intermediate that undergoes cyclization with concomitant release of the final cyclized product. Only the desired product is released in the CC step, while any side product or remaining intermediate cannot cyclize and thus remains bound to the solid phase this significantly eases the purification-work-up procedures to obtain the pure recovered compound from SPS. Several examples of CC are shown in Fig. 1.17 (1.47-1.50) and 1.18 (1.51-1.54). 

Where possible, enzymatic methods of cleavage should be used rather than chemical ones the latter do not generally proceed in good yield, and they are often accompanied by side reactions. Table 5-3 summarises a selection of reagents that have been used for chemical cleavage of proteins (Fontana and Gross 1986 Smith 1994b). 

Asymmetric synthesis via chiral, nonracemic hydrazones have gained wide acceptance in the community of synthetic organic chemists. Some examples of asymmetric alkylations of acyclic, satu-rated and unsaturated cyclic ketones - and aldehydes, which are applicable in the natural products field, are shown in Scheme 55. Various methods of cleavage of the alkylated hydrazones are also illustrated in Scheme 55. 

Once a synthetic sequence has been completed, the N-protective Cb group can be removed either by hydrogenolysis or by treatment with hydrogen bromide in acetic acid. Discussion of relative merits of N-protective groups and methods of cleavage. ... 

Finally, disulphide bonds can be located by hydrolysing a protein to a mixture of peptides using either a proteinase or a specific chemical method of cleavage and the mixture can be analysed directly by fast-atom bombardment mass spectrometry (Chapter 3) and again after reduction of disulphide bonds (Yazdanparast et al., 1987). By identifying those peaks which disappear as a result of reduction and new peaks with appropriate masses that have taken their place, it is simple to assign disulphide bonds to the relevant amino-acid sequences. 

This type of Cj-extrusion reaction has been predominantly of theoretical interest, especially with respect to the correlation of carbene reactivity with the structure of the precursor and the method of cleavage/ However, some attempts to use this Cj-extrusion method to generate certain carbenes for synthetic uses have also been reported. Thus, hexamethoxycyclopropane (5) has been probed as a potential source of dimethoxycarbene (6). 

Method of cleavage Cleavage is generally performed by protodesilylation. The conditions depend very much on the nature of the aromatic system, as well as on the substitution. Either neat TFA, TFA vapor or TFA/CH2CI2 can be used. Electron-poor aromatic systems require treatment with CsF in DMF/ water (4 1) at 110 °C. 

Other classical methods of cleavage include acidic hydrolysis (eq. 1), reduction... 

Electrolysis, —1.5 V, L1C104, CH3OH, 90% yield. The conditions can be adjusted to form either the sulhde or disulhde. Other sections discussing the Troc group should be consulted for alternative methods of cleavage. 

Although this synthesis produces an essentially enantio-merically pure product, it hcis several drawbacks the difficulty of purifying the starting material, the need for sec-butyllithium to make the lithio derivative of and difficulties we encountered (22) at times in the methyl iodide cleavage. We decided therefore to look for yet another chiral oxathiane adjuvant - preferably one which would closely resemble the 4,4,6-trimethyloxathiane used in most of the preliminaury experiments shown in Table 1 - and for a better method of cleavage. 

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