• BMB Reports
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• 제37권4호
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• pp.422-428
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• 2004

Raw-starch-digesting $\alpha$-amylase (Amyl III) was purified to an electrophoretically pure state from the extract of a koji culture of Aspergillus awamori KT-11 using wheat bran in the medium. The purified Amyl III digested not only soluble starch but also raw corn starch. The major products from the raw starch using Amyl III were maltotriose and maltose, although a small amount of glucose was produced. Amyl III acted on all raw starch granules that it has been tested on. However, it was considered that the action mode of the Amyl III on starch granules was different from that of glucoamylase judging from the observation of granules under a scanning electron microscope before and after enzyme reaction, and also from the reaction products. Glucoamylase (GA I) was also isolated and it was purified to an electrophoretically pure state from the extract. It was found that the electron micrographic features of the granules after treatment with the enzymes were quite different. A synergistic effect of Amyl III and GA I was observed for the digestion of raw starch granules.

• Food Science and Biotechnology
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• 제17권1호
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• pp.180-183
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• 2008

Digestion properties of 3 types of cereals, white rice, brown rice, and barley, were measured after cooking or grinding. Regardless of the processing methods, white rice showed the highest rate and the greatest extent of digestion, whereas barley showed the lowest values. During the early digestion period, cooked white rice kernels had a larger k (kinetic constant) value than uncooked white rice flour, indicating that cooking induced faster digestion than grinding. In the case of brown rice and barley, the cell wall in cooked kernels remained intact and resulted in a lower k values than those of uncooked flour. However, after 3 hr of digestion, the total digestion extent was greater for the cooked brown rice and barley than that for uncooked flours. The high content of slowly digestible starch (SDS) in cooked brown rice and barley might be due to the starch fraction which was protected by the cell wall. The resistant starch (RS) content, however, was greater for the uncooked flours than that for cooked kernels. The cooked kernels of 3 cereal samples tested showed higher glycemic index (GI) values than the uncooked flours.

• 한국미생물생명공학회:학술대회논문집
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• 한국미생물생명공학회 2001년도 Proceedings of 2001 International Symposium
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• pp.40-45
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• 2001

Raw starch-digesting amylase (BF-2A, M.W. 93, 000 Da) from Bacillus circulans F-2 was converted to two components during digestion with subtilisin. Two components were separated and designated as BF-2A' (63, 000 Da) and BF-2B (30, 000 Da), respectively. BF-2A' exhibited the same hydrolysis curve for soluble starch as the original amylase (BF-2A). Moreover, the catalytic activities of original and modified enzymes were indistinguishable in $K_{m}$, Vmax for, and in their specific activity for soluble starch hydrolysis. However, its adsorbability and digestibility on raw starch was greatly decreased. Furthermore, the enzymatic action pattern on soluble starch was greatly different from that of the BF-2A. A smaller peptide (BF-2B) showed adsorb ability onto raw starch. By these results, it is suggested that the larger peptide (BF-2A') has a region responsible for the expression of the enzyme activity to hydrolyze soluble substrate, and the smaller peptide (BF-2B) plays a role on raw starch adsorption. A similar phenomenon is observed during limited proteinase K, thermolysin, and endopeptidase Glu-C proteolysis of the enzyme. Fragments resulting from proteolysis were characterized by immunoblotting with anti-RSDA. The proteolytic patterns resulting from proteinase K and subtilisin were the same, producing 63- and 30-kDa fragments. Similar patterns were obtained with endopeptidase Glu-C or thermolysin. All proteolytic digests contained a common, major 63-kDa fragment. Inactivation of RSDA activity results from splitting off the C-terminal domain. Hence, it seems probable that the protease sensitive locus is in a hinge region susceptible to cleavage. Extracellular enzymes immunoreactive toward anti-RSDA were detected through whole bacterial cultivation. Proteins of sizes 93-, 75-, 63-, 55-, 38-, and 31-kDa were immunologically identical to RSDA. Of these, the 75-kDa and 63-kDa proteins correspond to the major products of proteolysis with Glu-C and thermolysin. These results postulated that enzyme heterogeneity of the raw starch-hydrolysis system might arise from the endogeneous proteolytic activity of the bacterium. Truncated forms of rsda, in which the gene sequence encoding the conserved domain had been deleted, directed the synthesis of a functional amylase that did not bind to raw starch. This indicates that the conserved region of RSDA constitutes a raw starch-binding domain, which is distinct from the active centre. The possible role of this substrate-binding region is discussed.d.

• 한국식품영양학회지
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• 제36권5호
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• pp.368-378
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• 2023

Changes in contents of free sugars, amino acids, and fatty acids of legumes were analyzed for each phase of in vitro digestion. In addition, contents of resistant starch in raw and digested pulses were compared. Soybeans, kidney beans, cowpeas, and chickpeas were analyzed. An in vitro digestion model was used to analyze contents of nutrients using LC-MS and GC-MS. Stachyose in kidneybean, cowpea, and chickpea increased as the digestion phase progressed. In four types of legumes, raffinose slightly decreased or showed no significant difference between the Oral phase and the BBMV phase. Content of glucose, a monosaccharide, increased during the BBMV phase. During the digestion phase, levels of free amino acids and free fatty acids also increased. Content of resistant starch was reduced compared to that in the raw material. It was 0.01g/100 g food in soybean, 1.06 g/100 g food in red kidney bean, 0.77g/ 100g food in cowpea, and 0.76 g/100 g food in chickpea. It was confirmed that nutrients in the in vitro digestion model were liberated at each digestion phase with changes in the content of resistant starch. These results are expected to be used as fundamental data for obtaining bioavailability of nutrients.

• 한국미생물·생명공학회지
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• 제16권3호
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• pp.231-237
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• 1988

Tapioca 전분으로부터 ethanol을 생산하기 위한 여러 방법을 비교한 결과 extrudate를 이용하는 것이 460.5ι/ton의 alcohol 수율을 보여 가장 효과적이었다. Rhizopus sp. glucoamylase로 69시간 반응시켰을 때 extrudate로부터는 108.7mg/$m\ell$, 생 tapioca chip에서는 43.8mg/$m\ell$의 환원당이 생성되어 생전분보다 extrudate에 대한 glucoamylase의 분해력과 분해속도가 월등함을 알 수 있었다. Extrudate를 이용한 alcohol 발효를 수행한 결과 발효 초기에 alcohol 농도가 높아져 세균 오염방지 및 발효시간을 단축시키는 효과를 얻을 수 있었으나 extrusion 온도가 alcohol 발효에 미치는 영향은 크지 않았다. Glucoamylase가 생tapioca 보다는 extrudate에 더 용이하게 작용할 수 있는 것은 전분입자의 구조적 차이 때문임을 전자현미경으로 확인하였으며 extrudate와 생전분에 대한 분해력은 Rhizopus sp. glucoamylase 가 Aspergillus usamii glucoamylase보다 높았다.

• BMB Reports
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• 제37권4호
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• pp.429-438
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• 2004

Complementary DNAs encoding $\alpha$-amylases (Amyl I, Amyl III) and glucoamylase (GA I) were cloned from Aspergillus awamori KT-11 and their nucleotide sequences were determined. The sequence of Amyl III that was a raw starch digesting $\alpha$-amylase was found to consist of a 1,902 bp open reading frame encoding 634 amino acids. The signal peptide of the enzyme was composed of 21 amino acids. On the other hand, the sequence of Amyl I, which cannot act on raw starch, consisted of a 1,500 bp ORF encoding 499 amino acids. The signal peptide of the enzyme was composed of 21 amino acids. The sequence of GA I consisted of a 1,920 bp ORF that encoded 639 amino acids. The signal peptide was composed of 24 amino acids. The amino acid sequence of Amyl III from the N-terminus to the amino acid number 499 showed 63.3% homology with Amyl I. However, the amino acid sequence from the amino acid number 501 to C-terminus, including the raw-starch-affinity site and the TS region rich in threonine and serine, showed 66.9% homology with GA I.

• Journal of Microbiology and Biotechnology
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• 제2권3호
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• pp.189-196
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• 1992

A $\alpha$-l, 4-D-glucan maltohydrolase $(\beta$-amylase), secreted by the mesophilic aerobic bacterium Bacillus polymyxa No.26, was purified and characterized. The enzyme production was increased after a logarithmic phase of bacterial growth and paralleled with the onset of bacterial sporulation. By applying anion exchange chromatography and gel filtration the enzyme was purified 16.7-fold and had a specific activity of 285.7 units/mg. Two enzyme activities were eluted on a column of DEAE-Sephadex chromatography, and they were designated as E-I for a major enzyme peak and E-II for a minor peak. Of them, E-I enzyme peak was further purified by using gel chromatography. The molecular mass of this enzyme was determined to be 64, 000 daltons and consisted of a single subunit, showing an isoelectric point of 8.9. The enzyme was able to attack specifically the $\alpha$-l, 4-glycosidic linkages in soluble starch and caused its complete hydrolysis to maltose and $\beta$-limited dextrin. This amylolytic enzyme displayed a temperature optimum at $45^\circ{C}$ and a pH optimum at 7.0. The amino acid composition of the purified enzyme was quite similar to the other bacterial $\beta$-amylases reported. Surprisingly, the purified enzyme from this aerobe only exhibited hydrolytic activity on soluble starch, not on starch granules. The degradation of from starch by $\beta$-amylase was greatly stimulated by pullulanase addition. These results differentiated from other $\beta$-amylases reported. Based on a previous result that showed the enzyme system involves in effective degradation of raw starch granules, this result strongly suggested that the purified enzyme (E-I) can be a synergistic part of starch granule-digestion and E-II plays a crucial role in digestion of starch granules.

• Nutrition Research and Practice
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• 제3권2호
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• pp.149-155
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• 2009

In this study, we investigated the heat effect of digestion-resistant fraction (RF) from soybean on retarding bile acid transport in vitro. The RFs from soybean retarded bile acid transport. A raw, unheated RF of soybean (RRF-SOY) was significantly more effective than the heated RF of soybean (HRF-SOY). The RS1 which physically trapped in milled grains and inaccessible to digestive enzyme after 18 hrs incubation level of content in RRF-SOY was found to be as high as 24.1% and after heating the RS1 of HRF-SOY was significantly reduced to 16.8%. The X-ray diffraction pattern of RF from soybean was altered after heat treatment. The RFs from soybean were characterized by peak at diffraction angles of $12.0^{\circ}$ and $20.0^{\circ}$ corresponding to RS content. Cellulose contents of RRF-SOY was 5% higher than that of HRF-SOY and pentosan contents of RRF-SOY was 5% higher than that of HRF-SOY, too. Whereas the hemicellulose content of RRF-SOY was 13% lower than HRF-SOY.

• 한국미생물·생명공학회지
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• 제13권4호
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• pp.329-337
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• 1985

토양등으로부터 생전분 분해효소 생산능이 있는 409주의 곰팡이와 62주의 세균 및 2주의 방선균을 분리하고 이들 중에서 7-BU 균주를 선정하여 동정하고 밀기울을 기본배지로 한 효소 생산조건 및 생전분을 이용한 알코올 발효 실험 등을 실시하여 다음과 같은 결과를 얻었다. 1. 생전분 분해효소 생산능이 강하다고 인정된 7-BU 균주는 Rhizopus oryzae로 동정되었다 2. 7-BU 균주의 효소생산 최적온도는 3$0^{\circ}C$이고 pH는 4.0이었으며 배양 72시간 후에 최고의 역가를 보였고 밀기울 배지에 옥수수전분을 0.5% 첨가하였을 때 효소생산이 더욱 촉진되었다. 3. 조효소액에 의한 각종 생전분의 가수분해율은 반응 4일에 쌀전분 89.2%, 옥수수전분 85%, 고구마전분 82%, 감자전분 36%이었고 쌀가루는 86%, 옥수수가루 81%. 고구마가루 79%, 감자가루 30%이었다. 4. 옥수수가루, 쌀가루, 고구마가루와 감자가루를 원료로 하여 조효소액과 효모를 첨가한 후 알코올 발효 실험을 한 결과 담금 4일 후 쌀가루구 9.4%, 옥수수가루구 9.0%, 고구마가루구 8. 1%, 감자가루구 5.4%의 에탄을 생성을 보였다.

• 한국미생물·생명공학회지
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• 제10권2호
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• pp.123-132
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• 1982

Bacillus circulans F-2의 정제$\alpha$-amylase(F-2A)의 효소적특성과 각종 생전분의 분해율을 검토하고 다음과 같은 결과를 얻었다. 1. 분자양은 93,000, 등전점은 pH5.0부근, 최적pH는 6.0~6.5, 안정 pH범위는 5.5~12.0, 최적온도는 6$0^{\circ}C$, 4$0^{\circ}C$ 이하에서 안정하였다. 2. Mn$^{＋＋}$, Co$^{＋＋}$ 은 부활제로써 작용하였으나 Ag$^{＋}$, Hg$^{＋＋}$, Cu$^{＋＋}$은 저해제로써 작용하였다. 3, 본 효소의 활성에 SH기는 필수적이 아닌 것으로 생각된다. 4. Michaelis constant (Km)는 1.704 mg/$m\ell$ 이고 활성화에너지는 25~4$0^{\circ}C$ 에서 12.297 kcal/mole, 40~6$0^{\circ}C$ 에서 7.831kca1/mo1e이었다. 5. Soluble starch, Amylose, Amylopectin으로부터 생성되는 초기분해산물은 G$_{6}$이며 분해가 진행됨에 따라 G$_4$와 G$_2$가 생성되었다. 6. 각종 Oligosaccharide로부터 생성되는 분해산물은 다음과 같다. G$_4$$\longrightarrow$ G$_2$＋ G$_2$,G$_3$ ＋G$_1$,G$_{5}$$\longrightarrow$ G$_4$＋G$_1$,G$_{6}$$\longrightarrow$G$_4$＋ G$_2$,G$_{7}$ G$_4$,G$_{8}$$\longrightarrow$ G$_4$＋G$_4$, 7. 감자 생전분, Sago 생전분, Yam 생전분에 대하여 본효소는 Porcine pancreatic amylase와 Streptococcus bovis amylase보다 월등하게 높은 분해율을 나타내었다.