• Journal of Microbiology and Biotechnology
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• 제19권4호
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• pp.358-361
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• 2009

The purified endochitosanase(Mw 41 kDa) from bacterium Bacillus cereus D-11 hydrolyzed chitooligomers $(GlcN)_{5-7}$ into chitobiose, chitotriose, and chitotetraose as the final products. The minimal size of the oligosaccharides for enzymatic hydrolysis was a pentamer. To further investigate the cleavage pattern of this enzyme, chitooligosaccharide alcohols were prepared as substrates and the end products of hydrolysis were analyzed by TLC and HPLC. The chitosanase split $(GlcN)_4GlcNOH$ into $(GlcN)_3+(GlcN)_1GlcNOH$, and $(GlcN)_5GIcNOH$ into $(GlcN)_4+(GlcN)_1GlcNOH$ and $(GlcN)_3+(GlcN)_2GlcNOH$. The heptamer $(GlcN)_6GlcNOH$ was split into $(GlcN)_5$ [thereafter hydrolyzed again into $(GlcN_3+(GlcN)2]+(GlcN)_1GlcNOH$, $(GlcN)_4+(GlcN)_2GlcNOH$, and $(GlcN)_3+(GlcN)_3GlcNOH$, whereas $(GlcN)_{1-3}GlcNOH$ was not hydrolyzed. The monomers GlcN and GIcNOH were never detected from the enzyme reaction. These results suggest that D-11 chitosanase recognizes three glucosamine residues in the minus position and simultaneously two residues in the plus position from the cleavage point.

• 한국식품과학회지
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• 제48권2호
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• pp.165-171
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• 2016

찰옥수수가루의 제빵 또는 제과 등 식품가공적성 향상을 위한 기초자료로서 활용하고자 열수 처리 및 효소 처리에 따른 물리 화학적 특성을 분석하였다. HMT 처리한 찰옥수수가루의 경우 $L^*$이 감소하고, $a^*$와 $b^*$이 증가하고 조단백질의 함량과 조지방함량이 감소하였다. 또한 HMT 처리 시 수분흡수지수가 낮아지고 수분용해지수가 높아짐에 따라 찰옥수수가루 내의 녹말의 호화현상으로 녹말 입자들의 치밀도가 낮아지고 비결정성부분이 많아지는 것으로 생각된다. HMT 처리 시 높은 열 처리로 인해 DSC를 통해 호화엔탈피(${\Delta}H$)의 감소, RVA를 통해 점도(viscosity)의 감소 및 XRD를 통해 결정성이 낮아짐을 알 수 있었다. 반면 ANN 또는 효소 처리 시 색도의 변화는 없었지만, 조단백질 및 조지방 함량의 변화가 있었고, DSC를 통해 확인한 호화엔탈피값은 대조군보다 약간 증가하거나 비슷하고, RVA를 통해 점도가 증가하는 것을 확인하였다. 또한 XRD 결과 전형적인 A-type의 결정성을 보였다. 전체적인 결과를 볼 때, HMT 처리는 찰옥수수가루 특성이 변화함에 따라 즉석식품으로 활용가능 할 것으로 생각되며 ANN에 의한 찰옥수수가루의 경우에는 점증제로서의 활용이 가능할 것으로 사료된다. 효소 처리한 경우 ANN 처리한 찰옥수수가루와 비교했을 때 큰 차이가 없으므로 효소 처리 가공보다는 ANN 처리하여 사용하는 것이 더 효과적이라고 생각된다.

• 대한환경공학회지
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• 제39권5호
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• pp.265-276
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• 2017

폐광산은 방치된 광미 등으로 인하여 주변환경에 복합적인 중금속 오염을 야기한다. 이를 방지하기 위하여 국내에서는 1990년대 중반부터 석회석 등의 안정화제와 복토를 이용한 안정화 공법을 기반으로 토양개량사업이 시행 중이다. 복원된 토양의 상부에서는 작물의 재배로 인해 중금속이 고정된 안정화 층이 지화학적 변화를 겪게 되며 이에 따른 중금속의 용출 및 이동이 가능하므로 토양개량사업을 마친 토양에 대한 질 평가 등의 사후 관리는 반드시 필요하다. 토양의 질 평가를 하기 위해서는 이화학적 분석 또는 생물학적인 분석을 개별적으로 하기보다는 이들을 결합한 종합적인 분석이 필요하며 이를 통해 토양의 생태기능상태(ecological functional state)를 평가할 수 있다. 본 연구에서는 대상시료로 경상북도 봉화군 풍정 광산, 전라남도 광양시 점동 광산, 충청남도 서산시 서성 광산 인근 안정화 처리 토양과 안정화 처리가 되지 않은 오염, 비오염 토양을 선정하였다. 토양의 이화학적 성질인 pH, CEC, LOI와 중금속 농도를 측정하였고, 미생물 효소활성도와 비소환원유전자를 정량하였다. 다변량 통계분석을 바탕으로 모든 데이터를 분석하여 토양 생태기능상태를 평가하였다. 안정화 심도 토양과 상부복토, 하부오염토 간의 상관관계를 확인한 결과, 안정화 심도 토양에서 중금속의 농도가 높게 측정되었다. 그리고 풍정광산에서는 안정화 처리 심도 토양이 하부오염토와 유사한 특성을, 점동, 서성 광산에서는 안정화 심도 토양이 상부복토와 유사한 특성을 나타내었는데 이는 점동, 서성 광산 주변 상부복토의 생태기능상태가 좋지 않을 수 있음을 시사한다.

• 한국식품저장유통학회지
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• 제10권1호
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• pp.80-88
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• 2003

대두단백 가수분해 산물의 맛과 향을 개선하기 위해 효소에 의한 가수분해 system을 확립하기 위하여 단백질 가수분해 패턴이 서로 다른 효소로 생산된 단백 분해산물의 가수분해도와 표면 소수도 등을 측정하였다. 이들 분해물의 pattern을 SDS 전기영동으로 조사하였고, 효소반응에 의한 단백질분해물의 관능검사를 실시하였다. 각 균주가 생산한 단백질 분해효소의 pH 변화에 따른 효소의 활성은 No. 16 효소(Bacillu megarerium Bl6)와 No. 4효소(Aspergillus oryzae M4)는 pH 7.0에서 No. 95효소(Bacillu subtilis YG 95)와 No. 5효소(Mucor circinelloides M5)는 8.0에서 가장 높은 효소반응 활성을 보였다. 또한 반응온도에 따른 효소활성의 크기는 4가지 효소 모두 45$^{\circ}C$에서 가장 높은 활성을 나타내었다. 대두단백질 분해물의 SDS 전기영동 pattern변화에서, Bacillus megaterium Bl6과 Mucor circinelloides M5의 효소 (No. 16, No. 5)는 반응 후 분자량이 비교적 큰 peptide가 많이 생성되었으며, 효소반응 3시간 경과 후에도 분자량 66KD의 peptide를 확인할 수 있었다. 반면에 Bacillus subtilis YG 95와 Aspergillus oryzae M4의 효소(No. 95, No. 4)는 분자량 15KD∼45KD 미만의 작은 분자량의 peptide 물질이 주로 생성되었으며, 반응 2시간 경과후에는 30KD 미만의 저분자 Peptide가 주로 생성되었다. 이와 같은 결과는 HPLC 분석 결과와 일치하였다. 가수분해가 진행됨에 따라서 SDS 표면소수도가 크게 저하되었으며, Aspergillus oryzae M4 효소의 분해물의 가수분해도가 가장 높았다. 관능검사 결과, No. 4(Aspergillus oryzae M4)와 No. 95(Bacillus subtilis YG 95) 가 강한 쓴맛을 나타내었다. 각각의 효소들을 조합해서 분해한 단백분해물을 관능검사한 결과, Aspergillus oryzae M4 효소와 조합된 것의 대두단백질 분해물이 비교적 강한 쓴맛을 나타내었다. 분해물의 단맛은 시료별로 큰 차이가 나타나지 않았으나 Bacillu megarerium Bl6과 Aspergillus oryzae M4를 조합하였을 때 상대적으로 단맛의 정도가 높게 나타났다. 따라서 이와 같은 효소특성을 이용하여 대두단백질을 가수분해를 하였을 때 다양한 단백질 분해물의 제조에 이용이 가능할 것으로 사료된다.

• BMB Reports
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• 제28권2호
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• pp.100-106
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• 1995

S-adenosylmethionine (SAM) synthetase was purified to homogeneity from soybean (Glycine max) axes. The enzyme was purified 216-fold with a 1.5% yield by ammonium sulfate fractionation, acetone fractionation, ion exchange chromatography with DEAE-sephacel, gel filtration with Sephacryl S-300, and afffinity chromatography with ATP-agarose. The enzyme activity reached a maximum 3 days after germination. SAM synthetase had a subunit molecular weight of 57,000 daltons from a silver stained single band on SDS-PAGE. The molecular weight of the enzyme was 110,000 daltons from Sephacryl S-300 gel filtration. The enzyme was composed of two identical subunits. The $K_m$ values of the enzyme for L-methionine and ATP were 1.81 and 1.53 mM, respectively. The enzymatic activity was not affected by polyamines, agmatine, or SAM analogues, but was inhibited by SAM. The inhibition pattern was showed non-competitive for L-methionine and uncompetitive for ATP. The activity of SAM synthetase was inhibited by thiol-blocking reagents. The enzyme was induced by treatment with $10^{-3}$ M putrescine at germination. Experimental data revealed a possible novel regulation mechanism of polyamine biosynthesis through several endogenous intermediates.

• Journal of Microbiology and Biotechnology
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• 제17권6호
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• pp.897-904
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• 2007

The aim of this paper is to discuss the methodology of our investigation of the dynamics of protein degradation and the total in situ protealytic activity in meso/eutrophic, eutrophic, and hypereutrophic freshwater environments. Analysis of the kinetics and rates of enzymatic release of amino acids in water samples preserved with sodium azide allows determination of the concentrations of labile proteins $(C_{LAB})$, and their half-life time $(T_{1/2})$. Moreover, it gives more realistic information on resultant activity in situ $(V_{T1/2})$ of ecto- and extracellular proteases that are responsible for the biological degradation of these compounds. Although the results provided by the proposed method are general y well correlated with those obtained by classical procedures, they better characterize the dynamics of protein degradation processes, especially in eutrophic or hypereutrophic lakes. In these environments, processes of protein decomposition occur mainly on the particles and depend primarily on a metabolic activity of seston-attached bacteria. The method was tested in three lakes. The different degree of eutrophication of these lakes was clearly demonstrated by the measured real proteolytic pattern and confirmed by conventional trophic state determinants.

• Journal of Microbiology and Biotechnology
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• 제18권8호
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• pp.1401-1407
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• 2008

The roles of conserved amino acid residues (Va1329-Ala330-Asn331-Glu332), constituting an extra sugar-binding space (ESBS) of Thermus maltogenic amylase (ThMA), were investigated by combinatorial saturation mutagenesis. Various ThMA mutants were firstly screened on the basis of starch hydrolyzing activity and their enzymatic properties were characterized in detail. Most of the ThMA variants showed remarkable decreases in their hydrolyzing activity, but their specificity against various substrates could be altered by mutagenesis. Unexpectedly, mutant H-16 (Gly-Leu-Val-Tyr) showed almost identical hydrolyzing and transglycosylation activities to wild type, whereas K-33 (Ser-Gly-Asp-Glu) showed an extremely low transglycosylation activity. Interestingly, K-33 produced glucose, maltose, and acarviosine from acarbose, whereas ThMA hydrolyzed acarbose to only glucose and acarviosine-glucose. These results propose that the substrate specificity, hydrolysis pattern, and transglycosylation activity of ThMA can be modulated by combinatorial mutations near the ESBS.

• 한국식품조리과학회지
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• 제9권3호
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• pp.181-186
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• 1993

Potato and carrot are the main sourses of vitamin C and vitamin A each. As a folk ramedy, potato-carrot mixtures have been used to cure a stomach ulcer in Korea. To investigate the effects of carrot juice on the browning of potato juice, we examined the brewing in various ratio of two juices. We also investigated the role of potato juice in the carotene oxidation of carrot juice. Delta "L" values of potato juice were abruptly decreased after 5 min. reaction and they were very different from the juice mixture of potato and carrot. Those containing higher ratio of potato were decreased greatly. In blanching treatment to eliminate the effect of the enzymes in potato and carrot, delta "L" values of cooked potato were decreased a little but those of cooked carrot were decreased greatly. To investigate the fact that the inhibitory effect of carrot juice in potato browning was due to the dilution of polyphenolics of potato juices by carrot juice mixing, we added H20 equivalent to carrot water content to potato juice. The diluted sample showed less decreasing pattern than nondiluted sample. We also added appropriate amount of ${\beta}$-carotene to the same samples. Delta "L" values of with added B-carotene were more slowly decreased than those of without added B-carotene.

• Journal of Microbiology and Biotechnology
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• 제14권2호
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• pp.373-378
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• 2004

The regulatory mechanism of methionine biosynthesis in Corynebacterium glutamicum was analyzed at the protein arid gene expression level. O-Acetylhomoserine sulfhydraylase (encoded by metY) was inhibited by 10 mM methionine to a residual activity of 10% level, whereas no such inhibition was found with cystathionine $\gamma$-synthase (encoded by metB) and cystathionine $\beta$-lyase (encoded by metC). The enzymatic activity of homoserine acetyltransferase (encoded by metX) was repressed to a residual activity of 25% level by 10 mM methionine which was added to the growth medium. Cystathionine $\gamma$-synthase and cystathionine $\beta$-lyase were also repressed by 10 mM methionine, but only to a residual activity of 50-70% level. O-Acetylhomoserine sulfhydrylase was very sensitive to repression by 10 mM methionine, showing residual activity of 13%. In addition, homoserine acetyltransferase was also repressed by 10 mM cysteine to 50% of its original activity. No repression of the enzymes by S-adenosyl methionine was observed. The pattern of repression by methionine indicated that the metB and aecD genes might be regulated by a common mechanism, while the metA and metY genes are differently regulated.

• Mass Spectrometry Letters
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• 제1권1호
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• pp.29-32
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• 2010

Cross reacting antibodies can cause an overestimation of the results of immunoassays. Therefore, alternative methods are needed for the accurate quantification of steroids. Gas chromatography combined with isotope-dilution mass spectrometry (GC-IDMS) is developed to quantify urinary active androgens, testosterone, epitestosterone and dihydrotestosterone, which are clinically relevant androgens to both hair-loss and prostate diseases. The method devised involves enzymatic hydrolysis with $\beta$-glucuronidase, solid-phase extraction, liquid-liquid extraction using methyl tert-butyl ether and subsequent conversion to pentafluorophenyldimethylsilyl-trimethylsilyl (flophemesyl-TMS) derivatives for sensitive and selective analysis in selected-ion monitoring mode. Flophemesyl-TMS derivatization not only eliminates matrix interference but also has a good peak resolution within a 6 min-run. A selective and sensitive GC technique with flophemesyl-TMS derivatives also allows accurate quantitative analysis of three active androgens when combined with IDMS. The limit of quantification of the three analytes was <50 pg/mL, and extraction recoveries ranged from 91.9 to 102.1%. The precision and accuracy were 1.2~6.5% and 89.0~106.7%, respectively. This GC-IDMS method can be useful for evaluating the drug efficacy and monitoring the biological processes responsible for male-pattern baldness and prostate diseases.