• Microbiology and Biotechnology Letters
• /
• v.28 no.4
• /
• pp.219-222
• /
• 2000

To produce and utilize microbial cyclomaltodextrinase being industrially useful, we isolated an alkalophilic Bacillus strain from soil which was capable of degrading cyclodextrins. The newly isolated strain was aerobic, gram-positive, spore-forming, motile, rod shape(0.2~0.4$\times$1.4~4.4 $\mu\textrm{m}$), and 35.8 mol% of DNA base composition. Based on its morphological, phisiological, and biochemical properties, it was identified as alkalophilic Bacillus sp. KJ-133 and cultivated well in the ranges of $30~40^{\circ}C$ and pH 8.0~9.0 . The cyclomaltodextrinase of the strain showed maximal production after 48h of cultivation at $37^{\circ}C$, and the activity was inhibited by Ag2+, Hg2+, Cu2+, and p-chloromercuribenzoate.

• Food Science and Biotechnology
• /
• v.18 no.3
• /
• pp.776-781
• /
• 2009

A putative cyclomaltodextrinase (BHCD) gene was found from the genome of Bacillus halodurans C-125, which encodes 578 amino acids with a predicted molecular mass of 67,279 Da. It shares 42-59% of amino acid sequence identity with common cyclomaltodextrinase (CDase)-family enzymes. The corresponding gene was cloned by polymerase chain reaction (PCR) and the dimeric enzyme with C-terminal 6-histidines was successfully overproduced and purified from recombinant Escherichia coli. BHCD showed the highest activity against ${\beta}-CD$ at pH 7.0 and $50^{\circ}C$. Due to its versatile hydrolysis and transglycosylation activities, BHCD has been confirmed as a member of CDases. However, BHCD can be distinguished from other typical CDases on the basis of its novel multisubstrate specificity. While typical CDases have over 10 times higher activity on ${\beta}-CD$ than starch or pullulan, the CD-hydrolyzing activity of BHCD is only 2.3 times higher than pullulan. In particular, it showed significantly higher activity ratio of maltotriose to acarbose than other common CDase-family enzymes.

• Microbiology and Biotechnology Letters
• /
• v.44 no.3
• /
• pp.363-369
• /
• 2016

A putative cyclomaltodextrinase gene (licd) was found from the genome of Listeria innocua ATCC 33090. The licd gene is located in the gene cluster involved in maltose/maltodextrin utilization, which consists of various genes encoding maltose phosphorylase and sugar ABC transporters. The structural gene encodes 591 amino acids with a predicted molecular mass of 68.6 kDa, which shares less than 58% of amino acid sequence identity with other known CDase family enzymes. The licd gene was cloned, and the dimeric enzyme with C-terminal six-histidines was successfully produced and purified from recombinant Escherichia coli. The enzyme showed the highest activity at pH 7.0 and 37℃. licd could hydrolyze β-cyclodextrin, starch, and maltotriose to mainly maltose, and it cleaved pullulan to panose. It could also catalyze the hydrolysis of acarbose to glucose and acarviosine-glucose. In particular, it showed significantly higher activity towards β-cyclodextrin and maltotriose than towards starch and acarbose. licd also showed transglycosylation activity, producing α-(1,6)- and/or α-(1,3)-linked transfer products from the acarbose donor and α-methyl glucopyranoside acceptor.

• Microbiology and Biotechnology Letters
• /
• v.41 no.4
• /
• pp.391-397
• /
• 2013

A putative cyclomaltodextrinase (LLCD) gene was cloned from the genome of Lactococcus lactis subsp. lactis KCTC 3769 (ATCC 19435), which encodes 584 amino acids with the predicted molecular mass of 68.7 kDa. KCTC 3769 shares approximately 40% of amino acid sequence identity with the CDase-family of enzymes. The dimeric enzyme with C-terminal six-histidines was heterologously expressed and purified from recombinant E. coli. LLCD showed the highest activity against ${\beta}$-cyclodextrin (CD) at pH 7.0 and $37^{\circ}C$. In particular, LLCD exhibited extremely low activity against starch and pullulan, while its CD-hydrolyzing activity was about 80 times higher than starch. Due to its much higher activity on CD over starch, LLCD has been identified as a member of CDases. However, LLCD can be distinguished from the other common CDases on the basis of its extremely low hydrolyzing activity against starch, pullulan, and acarbose.

• Korean Journal of Microbiology
• /
• v.53 no.3
• /
• pp.208-215
• /
• 2017

A cyclomaltodextrinase (SPCD) gene was cloned from Streptococcus pyogenes ATCC 700294. Its open reading frame consists of 567 amino acids (66.8 kDa), which shows less than 37% of amino acid sequence identity with the other CDase-family enzymes. The homo-dimeric SPCD with C-terminal six-histidines was expressed and purified from Escherichia coli. It showed the highest activity at pH 7.5 and $45^{\circ}C$, respectively. SPCD has the broad substrate specificities against ${\beta}$-cyclodextrin, starch, and maltotriose to produce mainly maltose, whereas it hydrolyzes pullulan to panose. It can also catalyze the hydrolysis of acarbose to glucose and acarviosine-glucose. Interestingly, it showed much higher activity on ${\beta}$-cyclodextrin and acarbose than that on starch, pullulan, or maltotriose, which makes SPCD distinguished from common CDase-family enzymes. Although SPCD has significantly high acarbose-hydrolyzing activity, it showed negligible transglycosylation activity.

• Proceedings of the Korean Society for Food Science of Animal Resources Conference
• /
• 2000.06a
• /
• pp.92-92
• /
• 2000

• Journal of Microbiology and Biotechnology
• /
• v.23 no.8
• /
• pp.1060-1069
• /
• 2013

Genome organization near cyclomaltodextrinases (CDases) was analyzed and compared for four different hyperthermophilic archaea: Thermococcus, Pyrococcus, Staphylothermus, and Thermofilum. A gene (CL1_0884) encoding a putative CDase from Thermococcus sp. CL1 (tccd) was cloned and expressed in Escherichia coli. TcCD was confirmed to be highly thermostable, with optimal activity at $85^{\circ}C$. The melting temperature of TcCD was determined to be $93^{\circ}C$ by both differential scanning calorimetry and differential scanning fluorimetry. A size-exclusion chromatography experiment showed that TcCD exists as a monomer. TcCD preferentially hydrolyzed ${\alpha}$-cyclodextrin (${\alpha}$-CD), and at the initial stage catalyzed a ring-opening reaction by cleaving one ${\alpha}$-1,4-glycosidic linkage of the CD ring to produce the corresponding single maltooligosaccharide. Furthermore, TcCD could hydrolyze branched CDs (G1-${\alpha}$-CD, G1-${\beta}$-CD, and G2-${\beta}$-CD) to yield significant amounts (45%, 40%, and 46%) of isomaltooligosaccharides (panose and $6^2$-${\alpha}$-maltosylmaltose) in addition to glucose and maltose. This enzyme is one of the most thermostable maltogenic amylases reported, and might be of potential value in the production of isomaltooligosaccharides in the food industry.

• Journal of Microbiology and Biotechnology
• /
• v.11 no.4
• /
• pp.712-715
• /
• 2001

A new combined method including the enzymatic hydrolysis of $\beta$-cyclodextrin ($\beta$-CD) and solvent extraction fo cholesterol from the hydrolyzed mixture was developed to recover cholesterol from a $\beta$-CD-cholesterol complex prepared from dairy products, such as cream, milk, and cheese. Cyclomaltodextrinase (cyclomatodextrin dextrin hydrolase, EC 3.2.1.54, DCase_ prepared form alkalophilic Bacillus sp. KJ 133 hydrolyzed the $\beta$-DC of the $\beta$-CD-cholesterol complex, and then, free cholesterol was efficiently extracted from the hydrolyzed mixture by a nonpolar solvent such as ethyl acetate. To increase the stability of free CDase, immobilized CDase was developed using sodium alginate as a carrier. The immobilized CDase showed a high recovery yield of cholesterol in a time-dependent manner compared to the free CDase. A gas chromatography analysis showed that more than 70% of cholesterol was recovered from the $\beta$-DC-cholesterol complex of cream by the immobilized CDase, whereas only 3% and 29% of cholesterol were recovered when the solvent extraction and free CDase treatment were used, respectively. The cholesterol recovered can be used as a raw material for steroid synthesis. Furthermore, this method can be an efficient way to recover cholesterol or other organic compounds that are bound in a $\beta$ -DC-cholesterol or -organic compound complex.

• Food Science and Biotechnology
• /
• v.15 no.6
• /
• pp.967-974
• /
• 2006

Cyclomaltodextrinases (CDases), maitogenic amylases, and neopullulanases share highly conserved primary structures and similar characteristics, and are thus classified into the same family. BLAST search has showed that a variety of bacterial strains harbor putative CDase family genes with several well-conserved motif amino acid sequences. In this study, four degenerate polymerase chain reaction (PCR) primer sets were designed for the detection of CDase genes, on the basis of their highly conserved amino acid blocks (WYQIFP, DGWRLD, LGSHDT, and KCMVW). The PCR detection conditions were optimized and the detection specificity of each for the primer sets was tested against the genomic DNAs isolated from 23 different Bacillus-associated species. Consequently, all tested primer sets evidenced successful amplification of specific PCR products in length, which share 55-98% amino acid sequence identity with known and putative CDases. The primers developed herein, therefore, can be applied for the easy and efficient detection and isolation of CDase family genes for the modification of functional food carbohydrates.

• The Microorganisms and Industry
• /
• v.27 no.1
• /
• pp.2-17
• /
• 2001

Cyclomaltodextrinase(CDase, EC 3.2.1.54), maltogenic amylase(EC 3.2.1.133). neopullulanase(EC 3.2.1.135)는 cyclomaltodextrin(CD), pullulan 및 전분을 가수분해하는 효소들이다. 이 효소들은 $\alpha$-1,4-Ο-glycosidic 결합에 작용하여 CD와 전분을 말토오스로 pullulan을 panose로 가수분해할 뿐만 아니라 올리고당들을 다양한 당 수용체 분자들의 C-3, C-4. C-6 수산기로 전이시키는 활성도 갖고 있다. 이러한 특성들은 기존의 $\alpha$-amylase를 비롯한 판수화물 분해효소들과 뚜렷이 구별되는 것으로 전분 분해효소들의 분류체계에 새로운 기준점을 제시한다고 하겠다. 본 총설에서는 CDase, maltogenic amylase, neopullulanase처럼 pullulan이나 전분보다 CD를 훨씬 더 잘 분해하는 효소들과 Thermoactinomyces vulgaris amylase II(TVA II)처럼 CD를 분해하기는 하나 pullulan을 더 잘 분해하는 효소들의 생화학적, 효소적, 구조적 특성들을 종합하여 소개하고자 하였다. 이 효소들은 40~60% 정도로 아미노산 서열이 동일하고, 세포 내에 존재하며, 분자량이 62~90 kDa로 $\alpha$-amylase보다 다소 크다. 아미노산 서열 비교분석 및 maltogenic amylase와 TVA II 등의 3차구조 분석 결과, 이 효소들은 아미노 말단에 보통 $\alpha$-amylase에는 존재하지 않는 약 130개 아미노산으로된 영역을 갖고 있어 이를 매개로 이합체를 형성할 수 있는 것으로 나타났다. 이합체-단위체 평형은 염 농도, 효소 농도, 산도 등에 의해 조절되고 단위체와 이합체 모두 효소환성을 갖고 있으나, 기질 특이성이 다르며 단위체는 전분을, 이합체는 CD를 선호하는데 이는 이합체 형성 시 활성부위의 구조적 변화에 따른 것으로 분석되었다. 본 총설에서는 CD 분해효소들의 다양한 기질 특이성을 올리고머 형성 등의 구조적 특성과 관련하여 논함으로써 관련 효소들의 분류체계를 보다 명확히 할 수 있는 자료를 제공하고자 하였으며, 이러한 효소들의 생리적 기능 및 산업적 이용에 대해 제안하고자 하였다.