Halophilic characterization of starch-binding domain from Kocuria varians α-amylase

Yamaguchi, Rui; Inoue, Yasuhiro; Tokunaga, Hiroko; Ishibashi, Matsujiro; Arakawa, Tsutomu; Sumitani, Jun-ichi; Kawaguchi, Takashi; Tokunaga, Masao

doi:10.1016/j.ijbiomac.2011.10.007

Cited by 13 publications

(20 citation statements)

References 34 publications

(65 reference statements)

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“…We have shown that SBD12 domain of KVA is the first example of halophilic CBM25 [34]. Consistent with its halophilic origin, the refolding rate of the SBD12 from thermal melting was enhanced with higher salt concentrations [34].…”

supporting

confidence: 62%

“…Several amylolitic enzymes, including Bacillus polymyxa and Bacillus circulans betaamylases, and B. circulans isocyclomaltooligosaccharide glucanotransferase, were reported to contain CBM25 member proteins [9,22,32]. We have shown that SBD12 domain of KVA is the first example of halophilic CBM25 [34]. Consistent with its halophilic origin, the refolding rate of the SBD12 from thermal melting was enhanced with higher salt concentrations [34].…”

mentioning

confidence: 59%

“…We have previously constructed expression plasmids pET15b-SBD12 and pET15b-SBD1, which encode SBD12 and SBD1 regions of KVA [34]. Expression vector, pET15b-SBD12, was transformed to E. coli BL21(DE3).…”

Section: Expression In and Purification From Escherichiamentioning

confidence: 99%

“…Harvested cells were sonicated in 50 mM Na-phosphate buffer, pH7.2, containing 0.15 M NaCl. Then, His-SBD12 protein was purified from this crude homogenate as reported previously [34]. His-SBD1 protein was expressed and purified as described above except for E. coli BL21(DE3) harboring pET15b-SBD1 plasmid.…”

Section: Expression In and Purification From Escherichiamentioning

confidence: 99%

“…In addition to the property described above, one of the most characteristic features that are shared by many halophilic proteins is the high solubility of the proteins both in the native and unfolded states [8,28,29,33,34]. High solubility of unfolded structure in aqueous buffer solution is the most critical parameter that plays a major role in its refolding efficiency.…”

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confidence: 99%

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Distinct Characteristics of Single Starch-Binding Domain SBD1 Derived from Tandem Domains SBD1-SBD2 of Halophilic Kocuria varians Alpha-Amylase

et al. 2012

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Kocuria varians alpha-amylase contains tandem starch-binding domains SBD1-SBD2 (SBD12) that possess typical halophilic characteristics. Recombinant tandem domains SBD12 and single domain SBD1, both with amino-terminal hexa-His tag, were expressed in and purified to homogeneity from Escherichia coli. The circular dichroism (CD) spectrum of His-SBD12 was characterized by a positive peak at 233 nm ascribed to the aromatic stacking. Although the signal occurred in the far UV region, it is an indication of tertiary structure folding. CD spectrum of single domain His-SBD1 exhibited the same peak position, signal intensity and spectral shape as those of His-SBD12, suggesting that the aromatic stacking must occur within the domain, and that two SBD domains in SBD12 and SBD1 has a similar folded structure. This structural observation was consistent with the biological activity that His-SBD1 showed binding activity against raw starch granules and amylose resin with 70-80% efficiency compared with binding of equimolar His-SBD12. Although the thermal unfolding rate of SBD12 and SBD1 were similar, the refolding rates of SBD12 and SBD1 from thermal melting were greatly different: His-SBD12 refolded slowly (T(1/2) = ~84 min), while refolding of single domain His-SBD1 was found to be 20-fold faster (T(1/2) = 4.2 min). The possible mechanism of this large difference in refolding rate was discussed. Maltose at 20 mM showed 5-6 °C increase in thermal melting of both His-SBD12 and His-SBD1, while its effects on the time course of unfolding and refolding were insignificant.

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supporting

confidence: 62%

mentioning

confidence: 59%

Section: Expression In and Purification From Escherichiamentioning

confidence: 99%

Section: Expression In and Purification From Escherichiamentioning

confidence: 99%

mentioning

confidence: 99%

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Distinct Characteristics of Single Starch-Binding Domain SBD1 Derived from Tandem Domains SBD1-SBD2 of Halophilic Kocuria varians Alpha-Amylase

et al. 2012

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Amyloid fibril formation in vitro from halophilic metal binding protein: Its high solubility and reversibility minimized formation of amorphous protein aggregations

et al. 2013

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Halophilic proteins are characterized by high net negative charges and relatively small fraction of hydrophobic amino acids, rendering them aggregation resistant. These properties are also shared by histidine-rich metal binding protein (HP) from moderate halophile, Chromohalobacter salexigens, used in this study. Here, we examined how halophilic proteins form amyloid fibrils in vitro. His-tagged HP, incubated at pH 2.0 and 58 C, readily formed amyloid fibrils, as observed by thioflavin fluorescence, CD spectra, and transmission or atomic force microscopies. Under these low-pH harsh conditions, however, His-HP was promptly hydrolyzed to smaller peptides most likely responsible for rapid formation of amyloid fibril. Three major acid-hydrolyzed peptides were isolated from fibrils and turned out to readily form fibrils. The synthetic peptides predicted to form fibrils in these peptide sequences by Waltz software also formed fibrils. Amyloid fibril was also readily formed from full-length His-HP when incubated with 10-20% 2,2,2-trifluoroethanol at pH 7.8 and 25 C without peptide bond cleavage.

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α-Amylase: an enzyme specificity found in various families of glycoside hydrolases

Janeček

Svensson

MacGregor³

2013

Cell. Mol. Life Sci.

300

234

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α-Amylase (EC 3.2.1.1) represents the best known amylolytic enzyme. It catalyzes the hydrolysis of α-1,4-glucosidic bonds in starch and related α-glucans. In general, the α-amylase is an enzyme with a broad substrate preference and product specificity. In the sequence-based classification system of all carbohydrate-active enzymes, it is one of the most frequently occurring glycoside hydrolases (GH). α-Amylase is the main representative of family GH13, but it is probably also present in the families GH57 and GH119, and possibly even in GH126. Family GH13, known generally as the main α-amylase family, forms clan GH-H together with families GH70 and GH77 that, however, contain no α-amylase. Within the family GH13, the α-amylase specificity is currently present in several subfamilies, such as GH13_1, 5, 6, 7, 15, 24, 27, 28, 36, 37, and, possibly in a few more that are not yet defined. The α-amylases classified in family GH13 employ a reaction mechanism giving retention of configuration, share 4-7 conserved sequence regions (CSRs) and catalytic machinery, and adopt the (β/α)8-barrel catalytic domain. Although the family GH57 α-amylases also employ the retaining reaction mechanism, they possess their own five CSRs and catalytic machinery, and adopt a (β/α)7-barrel fold. These family GH57 attributes are likely to be characteristic of α-amylases from the family GH119, too. With regard to family GH126, confirmation of the unambiguous presence of the α-amylase specificity may need more biochemical investigation because of an obvious, but unexpected, homology with inverting β-glucan-active hydrolases.

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Halophilic characterization of starch-binding domain from Kocuria varians α-amylase

Cited by 13 publications

References 34 publications

Distinct Characteristics of Single Starch-Binding Domain SBD1 Derived from Tandem Domains SBD1-SBD2 of Halophilic Kocuria varians Alpha-Amylase

Distinct Characteristics of Single Starch-Binding Domain SBD1 Derived from Tandem Domains SBD1-SBD2 of Halophilic Kocuria varians Alpha-Amylase

Amyloid fibril formation in vitro from halophilic metal binding protein: Its high solubility and reversibility minimized formation of amorphous protein aggregations

α-Amylase: an enzyme specificity found in various families of glycoside hydrolases

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