Histidine hydrolyzing

Histidine hydrochloride monohydrate L-Histidine Hydrolyzed gelatin Inositol Iron Iron ammonium citrate DL-lsoleucine L-lsoleucine L-Leucine ... 

A carboxylesterase (EC 3.1.1.1) from T. fusca [8, 86] and a steryl esterase (EC 3.1.1.13) from Melanocarpus albomyces [59] have also shown activity with PET oligomers and fabrics. The enzyme from M. albomyces with high specificity for fatty acid esters of sterols increased the hydrophilicity of PET fabrics. The highly hydrophobic serine hydrolase from T. fusca with a catalytic triad composed of serine, glutamic acid, and histidine hydrolyzed CTR and PET nanoparticles. The esterase showed high specificity towards short and middle chain-length fatty acyl esters of p-nitrophenol. In addition, p-nitrobenzyl esterases from Bacillus subtilis and B. licheniformis that hydrolyzed short chain dialkylphthalates and PET nanoparticles have been reported [74, 87]. 

Steps 3-4 of Figure 29.2 Hydrolysis The second nucleophilic acyl substitution step hydrolyzes the acyl enzyme and gives the free fatty acid by a mechanism analogous to that of the first two steps. Water is deprotonated by histidine to give hydroxide ion, which adds to the enzyme-bound acyl group. The tetrahedral... 

Kirk and Cohen (1969, 1971, 1973 Kirk et al., 1973) successfully applied the photo-fluoro-de-diazoniation method for the introduction of fluorine into imidazole and its derivatives with various substituents (e.g., histidine) 2- and 4-fluoroimi-dazole had not previously been synthesized by other methods. 2-Fluoroimidazole, in contrast to the 4-isomer, is easily hydrolyzed in water (Kirk et al., 1973). 

The second example of property space applications concerns the dipephde camo-sine (P-alanine-L-histidine, see Fig. 1.4) which represents the archetype of a series of histidine-containing dipeptides whose full physiological role remains poorly understood despite extensive studies in recent years [18-20]. Carnosine is synthesized by carnosine synthetase and hydrolyzed by dipeptidases (also called camosi-nases) which belong to the metalloproteases [21]. 

There are some side reactions that may occur when using EDC with proteins. In addition to reacting with carboxylates, EDC itself can form a stable complex with exposed sulfhydryl groups (Carraway and Triplett, 1970). Tyrosine residues can react with EDC, most likely through the phe-nolate ionized form of its side chain (Carraway and Koshland, 1968). The imidazolyl group of histidine may react with sulfo-NHS esters, resulting in an active carbonyl imidazole group which subsequently hydrolyzes (Cuatrecasas and Parikh, 1972). Finally, EDC may promote unwanted polymerization due to the usual abundance of both amines and carboxylates on protein molecules. 

As early as 1905 Abderhalden (Al) isolated from the hydrolyzate of the nondiffusible fraction of human urine four amino acids, i.e., leucine, alanine, glycine, and glutamic acid, and detected two others phenylalanine and aspartic acid. Some amino acids derived from this fraction have been quantitatively determined by Albanese et al. (A3) who found in the amount of the nondiffusible fraction corresponding to one liter of urine as much as 32.8 mg tryptophan, 18.0 mg phenylalanine, 16.2 mg methionine, 15.2 mg cystine, 13.1 mg arginine, 6.7 mg histidine, and 3.9 mg tyrosine. 

The mechanism by which serine peptidases, particularly serine endopep-tidases (EC 3.4.21), hydrolyze peptide bonds in peptides and proteins has been extensively investigated by X-ray crystallography, site-directed mutagenesis, detection of intermediates, chemical modification, H-NMR spectroscopy, and neutron diffraction [2-14], These studies revealed that all serine peptidases possess a catalytic triad, composed of a serine, a histidine, and an aspartate residue, and a so-called oxyanion hole formed by backbone NH groups. 

Eliminating deamination takes place in the degradation of histidine and serine. H2O is first eliminated here, yielding an unsaturated intermediate. In the case of serine, this intermediate is first rearranged into an imine (not shown), which is hydrolyzed in the second step into NH3 and pyruvate, with H2O being taken up. H2O does not therefore appear in the reaction equation. 

A potent enzyme inhibitor (abbreviated DEP) that acts by ethoxyformylation of proteins, usually at histidine residues. DEP is an irreversible inhibitor of ribonuclease, and rinsing glassware with a 0.1% (weight/volume) DEP solution is recommended to render glassware nuclease-free. Aqueous solutions must be freshly prepared for maximal effectiveness, because DEP will hydrolyze in 6-12 hours at neutral pH. 

The reaction proceeds as follows 1) An imine (Schiff base) is formed between a carboxyl group on the bound pyruvate and the amino group of histidine, with the elimination of a water molecule, 2) The bond to the carboxyl group of histidine is cleaved, liberating C02> 3) The second imine intermediate so formed is hydrolyzed to the free enzyme and histamine. 

To allow for the use of the stronger sulfating reagent pyridine/S03, an alternative protection strategy has been proposed by Fujii et al. 55-99 that is based on postsynthetic protection of the amino groups as Fmoc derivatives and of the serine hydroxy groups as terf-butyldiphenylsilyl derivatives as shown in Scheme 12. The histidine side chain is not protected since model studies indicate that sulfated histidine is readily hydrolyzed by water within 60 minutes. 

Aqueous surfactants are another class of catalysts. Substantial rate enhancement is seen in the reaction occurring at the micellar hydrocarbon-water interface, which is ascribed to a concentration of the reactant in the micellar pseudo-phase. Chiral p-nitrophenyl esters derived from phenylalanine are hydrolyzed by a histidine-containing dipeptide at a micellar interphase, at which a very high enantiomer discrimination, kR/ks up to 30.4 at 0°C, is observed (49). As shown in Scheme 20, the enantioselectivity is expressed at the stage at which a transient, zwitter-ionic tetrahedral intermediate leading to the acylimidazole is formed,... 

Nevertheless, malonyl-CoA is a major metabolite. It is an intermediate in fatty acid synthesis (see Fig. 17-12) and is formed in the peroxisomal P oxidation of odd chain-length dicarboxylic acids.703 Excess malonyl-CoA is decarboxylated in peroxisomes, and lack of the decarboxylase enzyme in mammals causes the lethal malonic aciduria.703 Some propionyl-CoA may also be metabolized by this pathway. The modified P oxidation sequence indicated on the left side of Fig. 17-3 is used in green plants and in many microorganisms. 3-Hydroxypropionyl-CoA is hydrolyzed to free P-hydroxypropionate, which is then oxidized to malonic semialdehyde and converted to acetyl-CoA by reactions that have not been completely described. Another possible pathway of propionate metabolism is the direct conversion to pyruvate via a oxidation into lactate, a mechanism that may be employed by some bacteria. Another route to lactate is through addition of water to acrylyl-CoA, the product of step a of Fig. 17-3. Tire water molecule adds in the "wrong way," the OH ion going to the a carbon instead of the P (Eq. 17-8). An enzyme with an active site similar to that of histidine ammonia-lyase (Eq. 14-48) could... 

PITC (phenylisothiocyanate) Aabs = 254 nm. Phenylthiocarbamyl amino acid derivatives are moderately stable at room temperature (1 day). PITC reacts well with both primary and secondary amino acids. Reaction time is approximately 5 minutes at room temperature. Excess reagent must subsequently be removed under vacuum. Also, for hydrolyzed samples, hydrochloric acid must be completely removed prior to derivatization. As a result, even though the actual reaction time is reasonably fast, the total time for various sample manipulations can add up to 2 hours. This is partially compensated by the extremely fast separation possible (12 minutes). Detection is by UV absorption only. Detection limits are typically in the high picomole range. Short column life can result due to unreacted PITC getting into the column. Unlike some of the other reagents, PITC quantifies tyrosine and histidine very well. PITC analysis is available as a commercially prepackaged system dubbed Pico-Tag by Waters Corporation. Representative references include 184-188. See Fig. 11 for a typical separation. 

P Felker. A gas-liquid chromatographic-isotope dilution analysis of cysteine, histidine, and tryptophan in acid-hydrolyzed protein. Anal Biochem 76 192-213, 1976. 

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