Free peptides

The procedure described is essentially that of Shioiri and Yamada. Diphenyl phosphorazidate is a useful and versatile reagent in organic synthesis. It has been used for racemlzatlon-free peptide syntheses, thiol ester synthesis, a modified Curtius reaction, an esterification of a-substituted carboxylic acld, formation of diketoplperazines, alkyl azide synthesis, phosphorylation of alcohols and amines,and polymerization of amino acids and peptides. - Furthermore, diphenyl phosphorazidate acts as a nitrene source and as a 1,3-dipole.An example in the ring contraction of cyclic ketones to form cycloalkanecarboxylic acids is presented in the next procedure, this volume. 

After the desired peptide has been made, treatment with anhydrous HF removes the final Boc group and cleaves the ester bond to the polymer, yielding the free peptide. 

A general step ahead in polycondensation was achieved by the application of the active ester method by DeTar et al.19) and Kovacs et al.291 Very soon, the nitrophenyl ester, the pentachlorophenyl ester, or the hydroxysucdnimido ester were used exclusively. The esters of the protected tripeptides could be purified by crystallization, then the N-protecting group was split off and the free peptide esters were purified again. Addition of base starts the polycondensation, resulting quickly in the formation of a viscous solution at low temperature. 

J Coste, E Frerot, P Jouin, B Castro. Oxybenzotriazole free peptide coupling reagents for N-methylated amino acids. Tetrahedron Lett 32, 1967, 1991. 

K Barlos, D Gatos, S Kutsogianni, G Papaphotiou, C Poulus, T Tsegenidis. Solid phase synthesis of partially protected and free peptides containing disulphide bonds by simultaneous cysteine oxidation-release from 2-chlorotrityl resin. Int J Pept Prot Res 38, 562, 1991. 

That the second term indeed can be neglected is illustrated in the following example. The CCR rates were measured in a small peptide weakly bound to STAT-6, a 100 kDa protein. The signals that are split by the heteronuclear j coupling are of equal intensity for the free peptide. After addition of protein, the signal intensities of the individual... 

The pH dependence of the trien reaction indicates that no protons are added before the rate-determining step. Hence, this step cannot come after the peptide bond breaks, because the metal-free peptide group must add a proton. Thus, the subsequent displacement reactions of the donors in the first chelate ring occur after the rate-determining step, which agrees with the lack of methyl group effects in this ring. 

There is some evidence for the population of a turn conformation similar to the bound conformation in the free peptide, however this is expected to be only one of many conformations adopted by the free peptide, as other NMR parameters (chemical shifts and values) indicated conformational... 

This enzyme [EC 3.4.19.1], also known as acylamino-acid releasing enzyme and A-acylpeptide hydrolase, catalyzes the hydrolysis of an acylaminoacyl-peptide to generate an acylamino acid and the free peptide. Catalysis is most... 

Serum antibodies were obtained by immunization with free peptide in complete Freund s adjuvant. 

Immunization with free peptides or with Intact Mb... 

Figure 9> Structure of the synthetic Mb peptides that were first used to study effect of peptide size on antibody response to immunization by free peptides (site 5, peptides 145-153 143-153 and 132-153) and to generate specific tolerance to preselected antigenic sites of Mb (peptides 143-153 135-153. 132-153. and truncated 139-153) The continuous lines in truncated peptide 139-153 indicate the absence of Ala and Tyr residues at positions 144 and 151. The residue in parenthesis is not required as part of the antigenic site in all antisera (1).

Other parameters of antibody response to free peptides. 

In another series of studies (57) we have found that there is an optimum dose for each immunizThg free peptide which varied with the peptide. It was also clear from these studies (57) that some free peptides are more imnunogenlc than others of equal size. For example, site 1 of Mb is more imnunogenlc than sites 2, 3, 4 or 5. At present the underlying reasons for differences in imnunogenlclty of free peptides of equal size are unknown. 

Having demonstrated that it Is possible to elicit antibodies with preselected submolecular binding specificities to protein regions by inmunization with free peptides (48,49,29,30,38,51), it appeared possible to produce monoclonal antibodies witlT similar predetermined specificities by the use of free synthetic peptides as imnunogens. The salient features in the development of... 

Table VI. Specificity of monoclonal antibodies from immunization with free peptide representing surface regions that are non-antigenic in whole Mb...

An interesting method for the selective persulfation of hydroxy groups in the presence of tyrosine has been proposed by Berg et al. 134 with the use of 100% H2S04/DMS/TFA (10 30 60) in the low acidity deprotection cocktail for resin-bound or free peptides with... 

According to this theoretical treatment, the slope of the plots of In k versus the solvent concentration, [3]m, can be employed to derive the contact area associated with the peptide-nonpolar ligand interaction. The retention and elution of a peptide in RPC can then be treated as a series of microequilibriums between the different components of the system, as represented by eq 6. The stoichiometric solvent displacement model addresses a set of considerations analogous to that of the preferential interaction model, but from a different empirical perspective. Thus, the affinity of the organic solvent for the free peptide P, in the mobile phase can be represented as follows ... 

Also free amino acids and peptides have been extracted from enamel. The material is very similar to that of dentine. Peptides from bone and teeth extract have an apparent molecular size of 750 to 5000 daltons163. Also peptides have been found in aqueous extracts of egg shells from birds and reptiles164 It has been assumed that free peptides and glycopeptides may have effects on the nucleation of the mineral phase185). On the other hand, those products could be split off from matrix proteins during crystallization processes. Perhaps there may be an association between free peptides and mucopolysaccharides or glycopeptides. It cannot be excluded that they are artefacts due to isolation procedures164). 

The syntheses of 4-[3-(trifluoromethyl)-3//-diazirin-3-yl]arene-containing peptides are carried out in a straightforward manner. In solid-phase syntheses, H-Phe(4-Tmd)-OH [Tmd = 3-(trifluoromethyl)-3//-diazirin-3-yl] is incorporated as the Fmoc-Phe(4-Tmd)-OH (Table 4).[i23,130-133] Solution-phase syntheses of [Phe4(4-Tmd)J-Leu-enkephalin and [Phe2(4-Tmd)J-aspartame employ Boc-Phe(4-Tmd)-OH and are carried out by the mixed anhydride method.1122 Apparently Phe(4-Tmd) is stable in TFA/CH2C12 (1 1) for 30 min at rt 122 or even in neat TFA for 15 min at 0°CJ134] A post-synthetic modification of a free peptide has been carried out by the mixed anhydride mediated coupling of 4-[3-(trifluoromethyl)-3/7-diazirin-... 

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