The polymorphism of barley β-amylase

Niku‐Paavola, Marja‐L; Skakoun, A.; Nummi, Martti; Daussant, J.

doi:10.1016/0005-2795(73)90191-8

Cited by 22 publications

(7 citation statements)

References 3 publications

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“…The stepwise decrease in Mr between 13-amylases I and 4 is consistent with the previously reported limited proteolysis in the COOHterminal region of[3-amylase 1 (34). Much wider ranges of size differences of cereal [3-amylases reported by other investigators are most likely due to intermolecular disulfide formation (42,43).…”

Section: Discussionsupporting

confidence: 88%

“…Ion exchange chromatography, isoelectric focusing and chromatofocusing thus revealed variable numbers of multiple forms (28,30,31,40) and a size range from Mr 40,000 to 400,000 (2,42,43) has been demonstrated by gel filtration, SDS-PAGE and ultracentrifugalion. Monomeric forms from barley were immunochemically identical, charge heterogeneous single polypeptide chains of Mr -60,000 (53).…”

Section: Introductionmentioning

confidence: 98%

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The four major forms of barley β-amylase. Purification, characterization and structural relationship

Lundgard

Svensson

1987

Carlsberg Res. Commun.

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Four major forms of barley 13-amylase have been purified in the presence of 0.1 M-thiol from extracts of flour fractionated by ammonium sulfate precipitation and chromatography on DEAE-cellulose and DEAE-Fractogel. The 13-amylases were NH2-terminally blocked, single polypeptide chains of approx. Mr 59,700, 58,000, 56,000 and 54,000, with corresponding pI 5.2, 5.3, 5.5 and 5.7. All forms displayed optimal activity on soluble starch between pH 4.5 and 7.5; all K m and Vmax values were 2.5 mg. ml -~ and 17 p.mol maltose, min -~ 9 (nmol protein) -t, respectively.The 13-amylase 4 consisted of different components terminating between Gly493 and Gin497 in the amino acid sequence deduced from the nucleotide sequence of a full length cDNA (KREIS et al., Eur. J. Biochem., in press (1987)). 13-Amylase 4 is the main product from limited proteolysis of the large form, [3-amylase 1, catalysed by malt protease(s), papain or subtilisin. Intermediate forms of ~-amylase with pl 5.3 and 5.5 were generated with crude malt enzymes, ct-chymotrypsin or thermolysin. The conversion effected by malt enzymes was prevented by leupeptin and EDTA in combination.

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Section: Discussionsupporting

confidence: 88%

Section: Introductionmentioning

confidence: 98%

The four major forms of barley β-amylase. Purification, characterization and structural relationship

Lundgard

Svensson

1987

Carlsberg Res. Commun.

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“…The present results show, however, that the bound ,3-amylase has a small but clear activity even on starch, but especially it rapidly hydrolyzes the small substrate, maltotetraose. Soluble polymers of,B-amylase are active (21) as well as the soluble complex of ,3-amylase and protein-Z (10). Thus, the attachment of ,B-amylase by S-S-bridges does not abolish its activity.…”

Section: Discussionmentioning

confidence: 99%

Release and Activity of Bound β-Amylase in a Germinating Barley Grain

Sopanen¹,

Laurière²

1989

Plant Physiol.

124

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In resting grains of Triumph barley (Hordeum vulgare L. cv Triumph) about 40% of the f-amylase could be extracted with a saline solution, the remaining 60% being in a bound form. During seedling growth (200C), the bound form was released mainly between days I and 3. When a preparation containing bound ,-amylase was incubated with an extract made of endosperms separated from germinating grains, release of bound ,#-amylase took place and could be studied in vitro. The release was almost completely prevented by leupeptin and antipain, specific inhibitors of a group of SH-proteinases, but it was not inhibited by pepstatin A or EDTA, which inhibit some other barley proteinases.It is thus very likely that in a whole grain, at least the bulk of the bound f-amylase is released by the proteolytic action of one or several SH-proteinases. When the bound jl-amylase was released by papain, its molecular weight was about 5000 daltons smaller than that of f8-amylase released by dithiothreitol. This indicates that the release is due to removal of a sequence of ,B-amylase itself. A similar decrease in size took place during seedling growth. Bound j0-amylase showed some activity against native starch and it hydrolyzed maltotetraose at a rate that was about 70% of the rate the same amount of bound jl-amylase gave after release. Bound j-amylase is thus not inactive and it is likely that the slower rate of hydrolysis is due to steric hindrances which prevent substrates from reaching the active site.In germinating barley grain, starch present in the starchy endosperm is hydrolyzed into glucose by a concerted action of a-and ,3-amylases, debranching enzyme and a-glucosidase (maltase) (2, 6). In contrast to the three other groups of enzymes, fl-amylase is not synthesized during germination but accumulates during development of the grain (9). At the end of development, when the grain is drying, a portion of the j3-amylase is attached, apparently via S-S-bridges, to insoluble constituents of the starchy endosperm (16,18 Our aim in the present study was to determine whether bound ,B-amylase is released in vivo by the action ofproteolysis and, if so, to identify the proteinase in question.Bound f3-amylase has often been cited to be latent (4, 14, 23). Here we show that it can hydrolyze different substrates with a rate depending on the size of the substrate. MATERIALS AND METHODS Plant MaterialGrains of barley (Hordeum vulgare L. cv Triumph) were obtained from SECOBRA (78580 MAULE, France). They were dehusked with 50% H2SO4, surface-sterilized with 1% NaOCl, and allowed to germinate aseptically on agar gel at 20°C in the dark (20). In these conditions the coleoptile was about 2 cm long after 3 d. Extraction of the Free and Bound ,8-AmylaseTwenty whole resting grains or endosperms from 20 germinating grains were homogenized at 20°C in a mortar with a small amount of quartz sand in the presence of 0.5 to 2 mL of 0.1 M NaCl. After homogenization, more NaCl solution was added, the total volume used being 8 mL. After centrifugation for 1...

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“…It catalyses the liberation of B-maltose from the nonreducing ends of starch and related 1,4-a-glucans. Most knowledge about this enzyme was acquired through studies of barley and bread wheat, and is essentially related to its genetic polymorphism (Joudrier and Bernard, 1977;Joudrier and Gobin, 1982;Ainsworth et al, 1983) or biochemical polymorphism (Nummi et al, 1965;Niku-Paavola et al, 1973;Lundgard and Svensson, 1987;Bureau et al, 1989;Gupta et al, 1991). The physiology of B-amylase was investigated during grain development (Kruger, 1972;Nishimura et al, 1987;Lam-&e et al, 1986) or germination (Daussant and Corvazier, 1970;Sopanen and Lam&e, 1989;Bureau et al, 1989;Guerin et al, 1992).…”

Section: Introductionmentioning

confidence: 99%

Heat Denaturation of Durum Wheat Semolina β‐Amylase Effects of Chemical Factors and Pasta Processing Conditions

Samson¹,

Morel²

1995

Journal of Food Science

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The susceptibility to heat denatnration of durum wheat @unylase was studied in aqueous solution at 50 to 65°C. The activation energy of the reaction of denatnration was 439 W/mole between 56 and 65°C. Additives were tested to determine their protective effects on enzyme activity (at 65°C) in aqueous solution. Maltose, resulting from degradation of starch by l3-amylase, appeared to be the most protective. The @amylase became more resistant to heat with higher concentrations of maltose, indicating an enzyme-maltose complex which was more stable to heat than the enzyme alone. The same mechanism occurred when durum wheat pasta was processed: maltose produced either during mixing and sheeting or during extrusion protected the enzyme. The degree of protection was proportional to the intensity of mechanical work imparted to the dough.

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The polymorphism of barley β-amylase

Cited by 22 publications

References 3 publications

The four major forms of barley β-amylase. Purification, characterization and structural relationship

The four major forms of barley β-amylase. Purification, characterization and structural relationship

Release and Activity of Bound β-Amylase in a Germinating Barley Grain

Heat Denaturation of Durum Wheat Semolina β‐Amylase Effects of Chemical Factors and Pasta Processing Conditions

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