Partial Purification of Acetic Acid Esterase from Malted Barley

Humberstone, F.J.; Briggs, D. E.

doi:10.1002/j.2050-0416.2002.tb00572.x

Cited by 3 publications

(5 citation statements)

References 22 publications

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“…By SDS-PAGE a single subunit protein band with an estimated M r of 19.7 kDa was observed (Figure 3a). The apparent molecular mass, determined under denaturing conditions, is comparable to the ones reported from the acetyl xylan esterases isolated from Pencillium purpurogenum (32), Bacillus pumilus (36) and acetic acid esterase from Barley malt (16). The estimated M r of acetic acid esterase under nondenaturing conditions using Sephacryl S-200 was found to be 79.4 kDa (Figure 3b), indicating it to be a homotetramer similar to the one reported from acetyl xylan esterase of Bacillus pumilus (36).…”

Section: Resultssupporting

confidence: 73%

“…were determined using 4-methylumbelliferyl acetate at 0.45 and 0.52 mM, respectively (38). A K m value of 25 mM for barley malt acetic acid esterase was reported using diacetin as the substrate (16). No information is available with respect to the detailed kinetics and substrate specificity of cereal/millet acetic acid esterases.…”

Section: Resultsmentioning

confidence: 99%

“…Acetic acid esterases obtained from plant and animal sources differ from microbial acetyl xylan esterases (EC 3.1.1.72) such as Trichoderma reesei, Aspergillus niger, and Schizophyllum commune (15) with respect to their substrate specificity. Partial characterization of acetic acid esterase from barley malt was recently reported (16).…”

Section: Introductionmentioning

confidence: 99%

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Purification and Partial Characterization of Acetic Acid Esterase from Malted Finger Millet (Eleusinecoracana,Indaf-15)

Latha

Muralikrishna

2007

J. Agric. Food Chem.

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Acetic acid esterase (EC 3.1.1.6) cleaves the acetyl groups substituted at O-2/O-3 of the xylan backbone of arabinoxylans and is known to modulate their functional properties. To date, this enzyme from cereals has not received much attention. In the present study, acetic acid esterase from 72 h ragi malt was isolated and purified to apparent homogeneity by a four-step purification, i.e., ammonium sulfate precipitation, DEAE-cellulose, Sephacryl S-200, and phenyl-Sepharose column chromatography, with a recovery of 0.36% and a fold purification of 34. The products liberated from alpha-NA and PNPA by the action of purified ragi acetic acid esterase were authenticated by ESI-MS and 1H NMR. The pH and temperature optima of the enzyme were found to be 7.5 and 45 degrees C, respectively. The enzyme is stable in the pH range of 6.0-9.0 and temperature range of 30-40 degrees C. The activation energy of the enzymatic reaction was found to be 7.29 kJ mol-1. The apparent Km and Vmax of the purified acetic acid esterase for alpha-NA were 0.04 microM and 0.175 microM min-1 mL-1, respectively. The molecular weight of the native enzyme was found to be 79.4 kDa by GPC whereas the denatured enzyme was found to be 19.7 kDa on SDS, indicating it to be a tetramer. EDTA, citric acid, and metal ions such as Fe+3 and Cu+2 increased the activity while Ni+2, Ca+2, Co+2, Ba+2, Mg+2, Mn+2, Zn+2, and Al+3 reduced the activity. Group-specific reagents such as eserine and PCMB at 25 mM concentration completely inhibited the enzyme while iodoacetamide did not have any effect. Eserine was found to be a competitive inhibitor.

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Section: Resultssupporting

confidence: 73%

Section: Resultsmentioning

confidence: 99%

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Purification and Partial Characterization of Acetic Acid Esterase from Malted Finger Millet (Eleusinecoracana,Indaf-15)

Latha

Muralikrishna

2007

J. Agric. Food Chem.

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“…The preparation obtained displayed esterase activity with pNPA and α-naphthyl acetate but also with acetylated xylan, confirming the proposal of Ward and Bamforth (2002) that the slow migrating esterases are AXE. Humberstone and Briggs (2002a) previously reported an acetic acid esterase from malt. The rate of thermal inactivation of esterase activity in this preparation was similar (Fig.…”

Section: Resultsmentioning

confidence: 99%

Barley Contains Two Cationic Acetylxylan Esterases and One Anionic Feruloyl Esterase

2005

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Cereal Chem. 82(6):621-625Various carbohydrate-active esterases are detected in extracts of malted barley when analyzed by polyacrylamide gel electophoresis. The slowest migrating and most heat-resistant of these are relatively cationic acetylxylan esterases. Two such activities, one with a high affinity for esterase substrates including acetylated xylan, and one with a low affinity, are indicated. These enzymes did not hydrolyze methyl ferulate. A relatively heat-labile anionic feruloyl esterase has also been purified. It has some, albeit low, ability to act on acetylated xylan. The feruloyl esterase effects extensive release of ferulate from endosperm cell walls isolated from barley, whereas the acetylxylan esterases are only capable of very limited release of acetate.

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“…Gel filtration was used as the third step in the purification. Previous gel filtration experiments used to purify acetyl esterase from barley malt 19 had shown that the best running buffer for the column contained both glucose (0.5M) and NaCl (2.5%). These additions had reduced interactions of the extract proteins with the column matrix.…”

Section: Urwhlqmentioning

confidence: 99%

Partial Purification of Ferulic Acid Esterase from Malted Barley

Humberstone¹,

Briggs²

2002

Journal of the Institute of Brewing

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A partial purification of ferulic acid esterase, which degrades feruloyl glycerol, has been achieved from barley malt in small yields. Coloured and viscous materials were removed from the malt extract using batch-elution anion exchange chromatography. Further steps included gradient elution anion exchange chromatography and gel filtration chromatography. Estimations of the molecular weight varied greatly from 22KDa to 158 KDa, possibly because the protein interacted with the matrix of the gel exclusion chromatography column and because multiple forms of the enzyme were present. The partially purified feruloyl esterase had an apparent Km of 0.46% feruloyl glycerol. However more than one enzyme may be present and the substrate contains two isomers and so Michelis-Menten kinetics may not be appropriate.

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Partial Purification of Acetic Acid Esterase from Malted Barley

Cited by 3 publications

References 22 publications

Purification and Partial Characterization of Acetic Acid Esterase from Malted Finger Millet (Eleusinecoracana,Indaf-15)

Purification and Partial Characterization of Acetic Acid Esterase from Malted Finger Millet (Eleusinecoracana,Indaf-15)

Barley Contains Two Cationic Acetylxylan Esterases and One Anionic Feruloyl Esterase

Partial Purification of Ferulic Acid Esterase from Malted Barley

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