P-nitrobenzyl Ester Research Articles

The interaction of alpha-chymotrypsin with phenylalanine derivatives containing a free alpha-amino group.

The action of α-chymotrypsin on L- and D-phenylalanine ethyl esters (PEE), L- and D-phenylalanine p-nitrobenzyl esters (PNBE), L-phenylalanine methyl and isopropyl esters, and N-methyl-L-phenylalamne methyl ester has been studied using a pH-stat. The D-esters were not hydrolyzed but acted as competitive inhibitors of the hydrolysis of the L-isomers. The N-methyl ester was very slowly hydrolyzed due to its low Kcat. For L-PEE (pK 7.23) and L-PNBE (pK 6.93), the activity of α-chymotrypsin is displaced to a more acid region relative to that for the N-acyl amino acid esters. The Km increases sharply below pH 6.5 while the kcat and kcat/Km show maxima at pH 6 and 7.6, respectively. On the acid side kcat is controlled by a basic group of pK 4.86 for L-PNBE and pK 5.1 for L-PEE, and kcat/Km by a basic group of pK 6.4 for L-PNBE and pK 6.6 for L-PEE. It is proposed that (i) deacylation is rate-limiting, (ii) in the catalytically active entities of the enzyme–substrate complex and acyl enzyme, the α-amino group of the substrate is protonated, (iii) the pK of the basic group on the acyl enzyme is considerably lowered by the presence of the [Formula: see text] of the substrate, and (iv) the increase in Km and Ki below pH 6.8 is due to the development of unfavorable charge interactions.

Antiradiation Compounds XIII: 1-(Dithioacetic Acid)-Pyridinium Betaines

Active mono-S-alkyl esters prepared from 1-(dithio-acetic acid)-pyridinium betaine (III) were found to be sufficiently stable for screening as radiation-protective agents, and e-withdrawing substituents in the pyridine ring gave stable betaines. Reaction of the methyl ester of III with phenacyl bromide and alkali resulted in S-alkylation to give a ketene mercaptal betaine (VIII). Both the allyl and p-nitrobenzyl esters of 1-(dithioacetic acid)-pyridinium halides were radiation-protective in mice, and betaines with substituents in the pyridine ring were radiation-protective in a bacterial test.

Insulin peptides. XVI. The synthesis of a nonapeptide and a dodecapeptide derivative related to the a chain of human insulin (positions 1-9 and 10-21).

Syntheses are described of protected peptides related to the N-terminus and C-terminus of the human insulin A chain. Thus the preparation is given of N-carbobenzox yglycyl-L-isoleucyl-L-valyl-7-f-butyl-L-glutamyl- L-glutaminyl-S-benzyl-L-cysteinyl-S-benzyl-L-cysteinyl-L-threonyl-L-serine hydrazide and N-carbobenzoxy-L-iso- leucyl-S-benzyl-L-cysteinyl-L-seryl-L-leucyl-L-tyrosyl-L-glutaminyl-L-leucyl-L-glutamyl- L- asparaginyl- L- tyrosyl - S - benzyl-L-cysteinyl-L-asparagine p-nitrobenzyl ester. The former derivative corresponds to positions 1-9 and the latter to positions 10—21 in the human insulin A chain sequence. R ecent communications3'4 from this laboratory described the synthesis of peptide derivatives related to the N-terminus of the A chain of sheep insulin and the synthesis of a dodecapeptide derivative con­ taining the C-terminal sequence of that chain. An in­ termediate in the construction of the latter peptide fragment, namely, the C-terminal nonapeptide portion, embodies the amino acid sequence found at the carboxyl terminus of the A chain of human insulin. The present report describes the synthesis of a protected nonapep­ tide and a protected dodecapeptide containing the N- terminal and C-terminal amino acid sequence, respec­ tively, of the human insulin A chain. The amino acid sequence of the reduced A chain of human insulin as determined by Nicol and Smith6 is glycylisoleucylvalylglutamylglutaminylcysteinylcyste- inylthreonylserylisoleucylcysteinylserylleucyltyrosylgul- taminylleucylglutamylasparaginyltyrosylcysteinylaspar- agine. An identical sequence was proposed for the reduced A chain of porcine insulin.6 The total synthesis of the human insulin A chain in the S-sul- fonated form is presented in the following paper7 and the preparation of two key intermediates used for that synthesis is described in the present report. One of these intermediates is the hydrazide of N-carboben- zoxyglycyl-L-isoleucyl-L-valyl - 7 - / - butyl -L - glutamyl - L - glutaminyl-S-benzyl-L-cysteinyl-S-benzyl-L-cysteinyl-L- threonyl-L-serine (I), the nonapeptide fragment which contains the amino acid sequence found at the amino terminus of the A chain, and the other intermediate is N - carbobenzoxy - L - isoleucyl - S - benzyl - L - cysteinyl - L - seryl-L - leucyl - L - tyrosyl - L - glutaminyl - L - leucyl - L - glu - tamyl-L-asparaginyl-L - tyrosy 1- S - benzyl - L - cysteinyl -L- asparagine ^-nitrobenzyl ester (XVI), the dodecapeptide

Chemistry of micrococcin P. Part XI. Application of a simple micromethod for the identification of lower volatile fatty acids to the structural study of 2-acylthiazoles

An acyl group attached to the 2-position of the thiazole ring is removed hydrolytically by hot mineral acid generating the related carboxylic acid which is identified as its p-nitrobenzyl ester by t.l.c. Direct chemical evidence has thus been obtained for the presence of a propionyl group in dimethyl micrococcinate. The procedure is applicable down to about the 0·5 µmole scale.

Use of diethyl chlorophosphite to acylate the hydroxy group of serine in peptides

1. Diethyl chlorophosphite “phosphorylates” the hydroxy group in serine, carbobenoxyserine and the p-nitrobenzyl ester of serine, and also in Gly-Ser and Ala-Ser. The reaction does not go to completion at room temperature, and instead a certain equilibrium is set up. 2. Diethyl chlorophosphite at room temperature does not acylate the N-terminal group of the amino acid or peptide.

The preparation of some benzyloxycarbonyl peptide p-nitrobenzyl esters derived from valine and glutamic acid

The preparation of three series of benzyloxycarbonyl peptide p-nitrobenzyl esters is described. In these derivatives a single L-valyl residue occupies different positions in a chain of γ-alkyl-L-glutamyl residues. The relative merits of various synthetic approaches to the compounds are also discussed.

P-Nitrobenzyl esters in the synthesis of peptides consisting of glycine and imino acids

1. By the mixed-anhydride and ethoxyacetylene methods some di-, tri-, tetra-, and hexa-peptides consisting of glycine, alanine, and imino acids were synthesized. 2. The advantages of using p-nitrobenzyl esters in peptide synthesis were demonstrated.