Action pattern and subsite mapping of <i>Bacillus licheniformis</i> α‐amylase (BLA) with modified maltooligosaccharide substrates

Kandra, Lili; Gyémánt, Gyöngyi; Remenyik, Judit; Hovánszki, György; Lipták, András

doi:10.1016/s0014-5793(02)02649-2

Cited by 53 publications

(58 citation statements)

References 28 publications

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“…Based on the cleavage pattern of a ρNP-maltoheptaoside, recently published for BLA (Kandra et al, 2002, and Fig. 2), and reports in the literature related to the maltosidase activities of various α-amylases, the production of G 2 ρNP and G 3 ρNP in addition to ρNP release is suggested.…”

Section: Methodsmentioning

confidence: 95%

Chemical Modification of Lysine Residues in Bacillus licheniformis α-Amylase: Conversion of an Endo- to an Exo-type Enzyme

Ebrahim‐Habibi

Khajeh²,

Nemat‐Gorgani³

2004

BMB Reports

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The lysine residues of Bacillus licheniformis α-amylase (BLA) were chemically modified using citraconic anhydride or succinic anhydride. Modification caused fundamental changes in the enzymes specificity, as indicated by a dramatic increase in maltosidase and a reduction in amylase activity. These changes in substrate specificity were found to coincide with a change in the cleavage pattern of the substrates and with a conversion of the native endo-form of the enzyme to a modified exo-form. Progressive increases in the productions of ρ-nitrophenol or glucose, when para nitrophenyl-maltoheptaoside or soluble starch, respectively, was used as substrate, were observed upon modification. The described changes were affected by the size of incorporated modified reagent: citraconic anhydride was more effective than succinic anhydride. Reasons for the observed changes are discussed and reasons for the effectivenesses of chemical modifications for tailoring enzyme specificities are suggested.

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Section: Methodsmentioning

confidence: 95%

Chemical Modification of Lysine Residues in Bacillus licheniformis α-Amylase: Conversion of an Endo- to an Exo-type Enzyme

Ebrahim‐Habibi

Khajeh²,

Nemat‐Gorgani³

2004

BMB Reports

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“…The low -limit dextrin content of the samples having low branch point density is in accordance with the specific action of -amylase on linear -1,4 chains. A previous study (Kandra, Gyémánt, Remenyik, Hovánszki and Lipták, 2002) demonstrated that the maximum frequency of α-amylase attack site is shifted towards the reducing end with longer chain length. Therefore, our data do not exclude that the distance between each branch points in the products were shorter or equal to four glucose units (Fig.…”

Section: α-Limit Dextrin Structure and β-Amylolysis Limitmentioning

confidence: 91%

Structure of branching enzyme- and amylomaltase modified starch produced from well-defined amylose to amylopectin substrates

Sorndech¹,

Sagnelli

Meier

et al. 2016

Carbohydrate Polymers

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Highlights •Enzyme modification of starch systems with well-defined amylose:amylopectin ratios provides detailed insight into the effects of substrate branching on enzymatic chain transfer • High amylose substrates produce high branching rates, high M w, short chains and low amylolytic digestion when treated with branching enzyme alone or in combination with amylomaltase.• Not only branching density but also molar mass of the glucan product restricts dietary degradation by steric hindrance towards human pancreatic α-amylase and α-glucosidases. AbstractThermostable branching enzyme (BE, EC 2.4.1.18) from Rhodothermus obamensis in combination with amylomaltase (AM, EC 2.4.1.25) from Thermus thermophilus was used to modify starch structure exploring potentials to extensively increase the number of branch points in starch. Amylose is an important constituent in starch and the effect of amylose on enzyme catalysis was investigated using amylose-only barley starch (AO) and waxy maize starch (WX) in well-defined ratios. All products were analysed for amylopectin chain length distribution, α-1,6 glucosidic linkages content, molar mass distribution and digestibility by using rat intestinal α-glucosidases. For each enzyme treatment series, increased AO content resulted in a higher rate of α-1,6 glucosidic linkage formation but as an effect of the very low initial branching of the AO, the final content of α-1,6 glucosidic linkages was slightly lower as compared to the high amylopectin substrates. However, an increase specifically in short chains was produced at high AO levels. The molar mass distribution for the enzyme treated samples was lower as compared with substrate WX and AO, indicating the presence of hydrolytic activity as well as cyclisation of the substrate. For all samples, increased amylose substrate showed decreased α-and β-amylolysis. Surprisingly, hydrolysis with rat intestinal α-glucosidases was higher with increasing α-1,6 glucosidic linkage content and decreasing M w indicating that steric hindrance towards the α-glucosidases was directed by the molar mass rather that the branching density of the glucan per se. Our data demonstrate that a higher amylose content in the substrate starch efficiently produces α-1,6 glucosidic linkages and that the present of amylose generates a higher M w and more resistant product than amylopectin. The combination of BEAMBE provided somewhat more resistant α-glucan products as compared to BE alone.

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“…Action patterns on 4-nitrophenyl a-D D-maltopentaoside, -hexaoside, and -heptaoside of the best characterized plant a-amylases [10], the barley isozymes AMY1, AMY2 and AMY1 mutants, emphasized the relationship between subsite site structures and productive substrate binding modes [11][12][13][14]. Longer MOS, however, are needed for accurate analysis of affinities in bacterial and plant aamylases with P 9 subsites long binding sites [3,[7][8][9]15,16]. Here, subsite mapping of AMY1 (including mutants at outer subsites) and AMY2 using a unique DP 3-12 2-chloro-4-nitrophenyl (CNP) MOS series [4,16], highlights the dynamic substrate affinity landscape and provides a rationale for reorganisation of binding energies through protein engineering.…”

Section: Introductionmentioning

confidence: 99%

Mapping of barley α‐amylases and outer subsite mutants reveals dynamic high‐affinity subsites and barriers in the long substrate binding cleft

et al. 2006

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Subsite affinity maps of long substrate binding clefts in barley a-amylases, obtained using a series of maltooligosaccharides of degree of polymerization of 3-12, revealed unfavorable binding energies at the internal subsites À3 and À5 and at subsites À8 and +3/+4 defining these subsites as binding barriers. Barley a-amylase 1 mutants Y105A and T212Y at subsite À6 and +4 resulted in release or anchoring of bound substrate, thus modifying the affinities of other high-affinity subsites (À2 and +2) and barriers. The double mutant Y105A-T212Y displayed a hybrid subsite affinity profile, converting barriers to binding areas. These findings highlight the dynamic binding energy distribution and the versatility of long maltooligosaccharide derivatives in mapping extended binding clefts in a-amylases.

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Action pattern and subsite mapping of Bacillus licheniformis α‐amylase (BLA) with modified maltooligosaccharide substrates

Cited by 53 publications

References 28 publications

Chemical Modification of Lysine Residues in Bacillus licheniformis α-Amylase: Conversion of an Endo- to an Exo-type Enzyme

Chemical Modification of Lysine Residues in Bacillus licheniformis α-Amylase: Conversion of an Endo- to an Exo-type Enzyme

Structure of branching enzyme- and amylomaltase modified starch produced from well-defined amylose to amylopectin substrates

Mapping of barley α‐amylases and outer subsite mutants reveals dynamic high‐affinity subsites and barriers in the long substrate binding cleft

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