Aminolysis amino acid esters

Peptides can be formed by aminolysis of iV-aminoacyl-3,5-dimethylpyrazole with free amino acid esters 303 as shown in Table 5-5. 

Acrylic acid esterified with cross-linked hydroxymethyl polystyrene or Wang resin reacts smoothly with primary or secondary aliphatic amines at room temperature (Entries 1 and 2, Table 10.6). Only sterically demanding amines or amines of low nucleophilicity (anilines, a-amino acid esters) fail to add to polystyrene-bound acrylate. Support-bound acrylamides are less reactive than acrylic esters, and generally require heating to undergo addition with amines (Entries 4 and 5, Table 10.6). a, 3-Unsaturated esters with substituents in the 3-position (e.g. crotonates, Entry 3, Table 10.6) react significantly more slowly with nucleophiles than do acrylates. The examples in Table 10.6 also show that polystyrene-bound esters are rather stable towards aminolysis, and provide for robust attachment even in the presence of high concentrations of amines. Entry 10 in Table 10.6 is an example of the alkylation of a resin-bound amine with an electron-poor alkene to yield a fluorinated peptide mimetic. 

The rapid aminolysis of cobalt(llI)-chelated glycine esters in aprotic solvents (Scheme 10 N4 = (en)2 or trien, R = Me, Et, R = H, CHR"C02Et) could be of value in peptide synthesis. The cobalt atom acts as both an N-protecting and an activating group. The synthesis of the chelated amino acid esters has presented some difficulties.207 A recent paper208 describes the use of methyl trifluoromethanesulfonate for the alkylation of chelated amino acids using dry trimethyl phosphate... 

Recently a simplified process was developed for incorporating l-methionine directly into soy proteins during the papain-catalyzed hydrolysis (21). The covalent attachment of the amino acid requires a very high concentration of protein and occurs through the formation of an acyl-enzyme intermediate and its subsequent aminolysis by the methionine ester added in the medium. From a practical point of view, the main advantage of enzymatic incorporation of amino acids into food proteins, in comparison with chemical methods, probably lies in the fact that racemic amino acid esters such as D,L-methionine ethyl ester can be used since just the L-form of the racemate is used by the stereospecific proteases. On the other hand, papain-catalyzed polymerization of L-methio-nine, which may occur at low protein concentration (39), will result in a loss of methionine because of the formation of insoluble polyamino acid chains greater than 7 units long. 

Figure 1. A simplified representation of the possible process for the papain-catalyzed hydrolysis and aminolysis E, enzyme (papain) S, substrate (protein) ES, Michaelis complex ES, peptidyl enzyme N, nucleophile (amino acid ester) P and P2, products formed from S by hydrolysis ...

All of the A -Trt amino acid fluorides listed in Table 6 are soluble in diethyl ether, with the exception of Trt-pGlu-F.t The fluorides undergo aminolysis with amino acid esters and amides without loss of configuration, although relatively slowly due to the steric hindrance resulting from the bulky Trt group. For example, the reaction of Trt-pGlu-F with H-His-OMe and Trt-Ile-F with H-Val-OBzl required 30 minutes and 6 hours respectively.t ... 

The direct aminolysis of unactivated esters and lactones is particularly difficult with secondary amines, since even primary amines require temperatures higher than 200 Application of high pressure (8 kbar, 1 bar = 100 kPa) at lower temperatures (room temperature to 45 C) makes this reaction possible. " Sodium cyanide catalyzes this reaction at atmospheric pressure and it proceeds without racemization of amino acid esters. 

Aminolysis with amino acid ester - Amide... 

Aso et al. [95] studied a model system in order to obtain basic information on the mechanism of amino acid incorporation during an enzymatic modification reaction in the presence of papain. They found that the amino acid ester reacted as a nucleophile in the aminolysis of the acyl-enzyme intermediate to result in the formation of new peptides. Several proteases used in enzymatic peptide bond synthesis are known to form transitory acyl-enzyme intermediates during the hydrolysis of proteins. However, the acyl groups can be transferred to other nucleophiles (amino terminals of peptides or amino acids), synthesizing new peptide bonds [71]. With full knowledge of the above-mentioned facts, covalent amino acid enrichment of proteins can result in... 

The traditional method for generation of protected peptide fragments in concert with the Boc-benzyl strategy is saponification or transesterification to cleave the PAM anchor. However, such methods are used less often today because of certain incompatibilities, as well as the development of more efficient techniques [95]. Allyl, o-nitrobenzyl, phenacyl (also cleaved by photolysis) [109,110], and fluorenylmethyl (cleaved by secondary amines) [111-113] handles all offer orthogonal mechanisms of release. The Kaiser oxime-resin is also popular for the production of protected segments it is cleaved via aminolysis with an amino acid ester or transesterification with hydroxypiperidine (reduction of this ester provides the free carboxyl) [97,114]. 

Various methods have been used to cleave protected peptides from the resin, including hydrazinolysis, ammonolysis, or aminolysis using a suitable amino acid ester [48-52]. Probably the most useful procedure, however, is transesterification of the peptide-resin with hydroxypiperidine. This initially forms the hydroxypiperidine ester of the protected peptide, which, after treatment with zinc in HOAc, furnishes the corresponding free carboxylic acid. This method does not lead to epimerization at the C-terminal amino acid or to loss of acid-sensitive protecting groups. Cleavage yields are usually high. 

A few years after the first publication on acylamino acid thiophenyl esters [4] the peptide research team of the CIBA laboratories in Basel, led by Robert Schwyzer, described a systematic study of the aminolysis of methyl esters substituted with various electron-withdrawing groups [20]. From a series of esters examined with respect to their reaction rates in aminolysis cyanomethyl esters were selected as best suited for peptide synthesis. Cyanomethyl esters were readily prepared through the reaction of acylamino acid salts with chloroaceto-nitrile and they showed satisfactory rates in the acylation of amino acid esters... 

A chemoenzymatic approach has been employed to synthesize enantioneriched P-amino acid esters. The aza-Michael addition of benzyl amine to P-alkyl substituted enoates, under solvent-free conditions, and subsequent hpase-catalyzed resolution via enantioselective aminolysis afforded the corresponding p-amino esters in moderate yields (19-36% yield) and with high enantioselectivities (93-99% ee) [96]. 

In general, naturally occurring (IV-protected) L-amino acid esters of short-chain alcohols, such as methyl and ethyl esters are usually sufficiently reactive as acceptors to achieve reasonable reaction rates in enzymatic peptide synthesis via aminolysis. For less reactive (nonnatural) analogs, such as a-substituted [313] or D-configured amino acids [314], activated esters are recommended, among them, 2-haloethyl (e.g., 2-chloroethyl, trifluoroethyl) [315], p-nitrophenyl [316], or guanidinophenyl esters [317]. For the use of cyclic activated esters [5(47/)-oxazolones] see below. 

In contradiction to this, the protein biosynthesis proceeds via aminolysis of activated amino acids reactively bound to t-RNA, where the growing protein is liberated from the polymer RNA support (Fig. 3). This type of biosynthesis was imitated in the way of a drastically simplified model reaction, namely the peptide syntheses with solid phase bound activated amino acid esters [37, 38], which here are not the matter for discussion (Fig. 4). 

Aminolysis of methoxy carbene metal pentacarbonyl complexes of the chromium group yields amino carbene compounds. If the aminolysis is performed with amino acid esters, amino carbene derivatives of amino acids are obtained [162,163]. The metal carbene is relatively stable under a variety of conditions but can be removed with TFA. It may therefore be regarded as an amino protecting group in peptide synthesis. The metal amino carbene amino acid may be activated and coupled to other amino acid esters. Following this idea, Weiss and Fischer prepared various dipeptides, the tripeptide (OC)5Cr(Ph)-Ala-Ala-Ala-OMe 69 [162] and the tetrapeptide (OC)5Cr(Ph)-Gly-Gly-Pro-Gly-OMe 70 (Scheme 5.35) [163]. Treatment with cone. TFA at 20 °C for 10 min. removes the metal carbene group and furnishes the free peptide esters along with Cr(CO)g. Removal of the metal carbene is also possible with 80% acetic acid (80 °C, 30 min.), which leaves Boc... 

Apart from aminolysis and olefin metathesis the photoactivation of aminocar-bene complexes offers another nonconventional entry into peptide synthesis. Irradiation into the hypsochromic MLCT-band of chromium aminocarbenes such as 201 generates a ketene-like intermediate 204 that is trapped by amino acid esters such as 202 or 205 to produce dipeptides 203 or 206 after enantioselective protonation (Scheme 11.49) [106]. This photochemical protocol generally combines good yields with high diastereoselectivities and is especially attractive for the incorporation of a-alkyl a-amino acid esters into peptides that may be hampered in conventional peptide synthesis methodologies due to steric hindrance [106c]. 

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