Bitterness of Phenylalanine- and Tyrosine-containing Peptides

Ishibashi, Norio; Sadamori, Koji; Yamamoto, Osamu; Kanehisa, Hidenori; Kouge, Katsushige; Kikuchi, Eiichi; Okai, Hideo; Fukui, Sakuzô

doi:10.1080/00021369.1987.10868574

Cited by 44 publications

(56 citation statements)

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“…Exopeptidases can reduce the amount of peptides with undesirable tastes through the removal of a single hydrophobic amino acid, or pairs of them, from the terminal ends. For example, phenylalanine-containing peptides taste Ͼ100-fold more bitter than does free phenylalanine (13,14). Control and termination of the hydrolytic reaction are therefore crucial for obtaining hydrolysates with the desired organoleptic properties.…”

mentioning

confidence: 99%

Aminopeptidase C of Aspergillus niger Is a Novel Phenylalanine Aminopeptidase

Basten

Dekker

Schaap

2003

Appl Environ Microbiol

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A novel enzyme with a specific phenylalanine aminopeptidase activity (ApsC) from Aspergillus niger (CBS 120.49) has been characterized. The derived amino acid sequence is not similar to any previously characterized aminopeptidase sequence but does share similarity with some mammalian acyl-peptide hydrolase sequences. ApsC was found to be most active towards phenylalanine ␤-naphthylamide (F-␤NA) and phenylalanine para-nitroanilide (F-pNA), but it also displayed activity towards other amino acids with aromatic side chains coupled to ␤NA; other amino acids with nonaromatic side chains coupled to either pNA or ␤NA were not hydrolyzed or were poorly hydrolyzed. ApsC was not able to hydrolyze N-acetylalanine-pNA, a substrate for acyl-peptide hydrolases.Many food products contain flavors obtained by the hydrolysis of proteins. These peptides and amino acids can taste sweet, sour, or bitter. Mixtures of endoproteases are often deliberately used in conjunction with exoproteases to improve these food flavors. Exopeptidases can reduce the amount of peptides with undesirable tastes through the removal of a single hydrophobic amino acid, or pairs of them, from the terminal ends. For example, phenylalanine-containing peptides taste Ͼ100-fold more bitter than does free phenylalanine (13,14). Control and termination of the hydrolytic reaction are therefore crucial for obtaining hydrolysates with the desired organoleptic properties.Enzymes from Aspergillus niger have been used in food production for several decades, and five different endoproteases (PepA to PepE [see reference 26 and references therein]), two carboxypeptidases (CpdI and PepF/CpdII [9, 24, 25]), and one aminopeptidase (ApsA [4]) have been cloned and characterized. Experiments have shown that a particular enzyme preparation of A. niger that contains a specific aminopeptidase activity can liberate phenylalanine from proteins that are present in dough and (semi)hard cheeses (8), thereby improving the flavor and aroma of these products. So far, only one phenylalanine-specific aminopeptidase has been characterized, namely, APF1 from the basidiomycetous fungus Schizophyllum commune (6). APF1 is an intracellular zinc metallo-aminopeptidase that can also hydrolyze an N-terminal tyrosine.In this report, we describe a new aminopeptidase. The gene was cloned from A. niger, and purification and characterization of the gene product show that the gene encodes an aminopeptidase that specifically hydrolyzes amino-terminal phenylalanine and other amino acids with aromatic side groups. ). To study the possible regulation of apsC messenger levels by the carbon and nitrogen source, strain N402 was grown on MM supplemented with either 1% glucose, 1% fructose, and 0.6% NH 4 Cl or 0.4% NaNO 3 or on MM supplemented with 1% bovine serum albumin, 1% elastin, or 1% collagen. RNA was isolated from these cultures and subjected to Northern analysis. MATERIALS AND METHODS StrainsPurification of extracellular ApsC from A. niger NRRL 3112 and peptide sequencing. A. niger NRRL 3112 was grown in a...

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mentioning

confidence: 99%

Aminopeptidase C of Aspergillus niger Is a Novel Phenylalanine Aminopeptidase

Basten

Dekker

Schaap

2003

Appl Environ Microbiol

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“…Therefore, the enzymatic production of amino acid mixtures from proteins using a combination of proteases and peptidases has recently been paid considerable attention in the food protein processing industry. However, the enzymatic hydrolysis of proteins results in a bitter peptide taste, due to the formation of low molecular weight peptides composed mainly of hydrophobic amino acids [8][9][10]20]. Further hydrolysis of bitter peptides is carried out using aminopeptidase, alkaline/neutral protease and carboxypeptidase.…”

Section: Introductionmentioning

confidence: 99%

Purification and characterization of hyperthermotolerant leucine aminopeptidase from Geobacillus thermoleovorans 47b

Deejing¹,

Yoshimune²,

Lumyong³

et al. 2005

J IND MICROBIOL BIOTECHNOL

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A thermophilic bacterium, which we designated as Geobacillus thermoleovorans 47b was isolated from a hot spring in Beppu, Oita Prefecture, Japan, on the basis of its ability to grow on bitter peptides as a sole carbon and nitrogen source. The cell-free extract from G. thermoleovorans 47b contained leucine aminopeptidase (LAP; EC 3.4.11.10), which was purified 164-fold to homogeneity in seven steps, using ammonium sulfate fractionation followed by the column chromatography using DEAE-Toyopearl, hydroxyapatite, MonoQ and Superdex 200 PC gel filtration, followed again by MonoQ and hydroxyapatite. The enzyme was a single polypeptide with a molecular mass of 42,977.2 Da, as determined by matrix-assisted laser desorption ionization and time-of-flight mass spectrometry, and was found to be thermostable at 90 degrees C for up to 1 h. Its optimal pH and temperature were observed to be 7.6-7.8 and 60 degrees C, respectively, and it had high activity towards the substrates Leu-p-nitroanilide (p-NA)(100%), Arg-p-NA (56.3%) and LeuGlyGly (486%). The K(m) and V(max) values for Leu-p-NA and LeuGlyGly were 0.658 mM and 25.0 mM and 236.2 micromol min(-1) mg(-1) protein and 1,149 micromol min(-1) mg(-1) protein, respectively. The turnover rate (k(cat)) and catalytic efficiency (k(cat)/ K(m)) for Leu-p-NA and LeuGlyGly were 10,179 s(-1) and 49,543 s(-1) and 15,470 mM(-1 ) s(-1) and 1981.7 mM(-1 ) s(-1), respectively. The enzyme was strongly inhibited by EDTA, 1,10-phenanthroline, dithiothreitol, beta-mercaptoethanol, iodoacetate and bestatin; and its apoenzyme was found to be reactivated by Co(2+) .

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“…Additionally, the higher the contents of hydrophobic amino acids at the C-terminus, the higher the level of bitterness (Ishibashi et al 1987a, b;Kanehisa et al 1984;Otagiri et al 1984;Shinoda et al 1986). The tripeptides Phe-Phe-Phe and Tyr-Tyr-Tyr are bitter, and both of them are fivefold bitterer than caffeine (Ishibashi et al 1987b). In contrast, their corresponding dipeptides Phe-Phe and Tyr-Tyr have 83 and 43 % the bitterness of caffeine, respectively (Ishibashi et al 1987b).…”

Section: Profiles Of Casein Hydrolysate Hydrolyzed By Th2-cpmentioning

confidence: 99%

“…The tripeptides Phe-Phe-Phe and Tyr-Tyr-Tyr are bitter, and both of them are fivefold bitterer than caffeine (Ishibashi et al 1987b). In contrast, their corresponding dipeptides Phe-Phe and Tyr-Tyr have 83 and 43 % the bitterness of caffeine, respectively (Ishibashi et al 1987b). Among the synthetic peptides tested, Phe-Phe-Phe and Tyr-Tyr-Tyr were the optimum two substrates of TH2-CP (Fig.…”

Section: Profiles Of Casein Hydrolysate Hydrolyzed By Th2-cpmentioning

confidence: 99%

Characterization of a novel metallocarboxypeptidase from Streptomyces cinnamoneus TH-2

Wan¹,

Uraji²,

Arima

et al. 2016

Bioresour. Bioprocess.

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Background: Carboxypeptidases are exopeptidases that catalyze the release of amino acids from the C-terminus of peptides or proteins. The peptides consisting of hydrophobic amino acids taste bitter. Therefore, the hydrolytic capability of carboxypeptidase toward hydrophobic amino acids at the C-terminus of peptides is useful for the degradation of bitter peptides. Results:Using the genome data of Streptomyces cinnamoneus TH-2, we expressed and characterized a novel metallocarboxypeptidase (TH2-CP) in Streptomyces lividans. TH2-CP had a molecular mass of 37.7 kDa. As TH2-CP possesses a zinc-binding consensus motif (HXXE……H) and N-terminal prosegment residues, we suggest that TH2-CP could be classified into the M14A subfamily. In the presence of Z-Gly-Leu as the substrate, TH2-CP showed optimum activity at pHs 7 and 8 in potassium phosphate and Tris-HCl buffers, respectively. The optimum temperature for activity was 51 °C. Furthermore, 50 % activity was conserved after incubation at 38 °C for 30 min. TH2-CP showed broad substrate specificity, with a preference for hydrophobic amino acids, as demonstrated by casein hydrolysate breakdown. Conclusions:A novel metallocarboxypeptidase, TH2-CP, from S. cinnamoneus TH-2 was characterized. TH2-CP preferred substrates with hydrophobic amino acids at the C-terminal position for casein peptides. This property indicates that TH2-CP can be used to decrease the bitterness of peptides in food industries.

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Bitterness of Phenylalanine- and Tyrosine-containing Peptides

Cited by 44 publications

References 3 publications

Aminopeptidase C of Aspergillus niger Is a Novel Phenylalanine Aminopeptidase

Aminopeptidase C of Aspergillus niger Is a Novel Phenylalanine Aminopeptidase

Purification and characterization of hyperthermotolerant leucine aminopeptidase from Geobacillus thermoleovorans 47b

Characterization of a novel metallocarboxypeptidase from Streptomyces cinnamoneus TH-2

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