• 치위생과학회지
• /
• 제2권2호
• /
• pp.121-124
• /
• 2002

본 연구에서는 구강미생물에서 발견되는 FTF 효소의 hydroxyapatite에 대한 친화도와 상호간의 결합에 영향을 주는 요소들을 비교하였으며, hydroxyapatite에 결합된 FTF 효소의 효소활성은 매우 안정적으로 높은 효소활성을 보였다. 결과를 요약하면 다음과 같다. 1. FTF는 치아를 대신하여 사용한 hydroxyapatite의 표면상태와 표면적에 영향을 받는 것으로 나타났다. 2. Hydroxyapatite에 결합된 FTF는 높은 플락탄 생성능을 보여, 비결합 FTF와 비교시, 약 70%의 효소활성을 나타내었다. 3. 결합된 FTF는 비결합 FTF와 비교시, 단백질분해효소의 분해작용에 대한 높은 저항성을 보여, hydroxyapatite에 결합된 효소에서는 3차원적인 구조적 변화가 나타나는 것으로 판단된다.

• 미생물학회지
• /
• 제19권3호
• /
• pp.137-141
• /
• 1981

Occurrence and distribution of sucrose metabolizing enzymes in oral streptococci had been studied. In these studies, the carbohydrate component of the culture medium had been glucose. I have extended these studies by analyzing bacterial culture supernatants for the relative content of hexosyltransferases, namely glucosyl and fructosyltransferase. As a carbohydrate, fructose was used. The growth measured for nine oral streptococci (Strptococcus mutans strains BHT, ING, AHT, 6715, LM-7, and SL-1 ; Streptococcus sanguis 903, 9811, and M-5) varied. The level of glucosyltansferase activity also varied among S. mutans strains, and its level in S. sanguis was relatively low. Fructosyltansferase activity of the various strains fluctuated more than of glucosyltransferase. S.mutans strain LM-7 had significantly higher level of both enzymes. As a whole, fructose-grown cultures had generally an agreeable trend of enzyme activity to those from glucose-grown cultures.

• 한국생물공학회:학술대회논문집
• /
• 한국생물공학회 2002년도 생물공학의 동향 (X)
• /
• pp.239-242
• /
• 2002

We have synthesized branched oligosaccharides (BOS) by the mixed-culture fermentation (MBOS), fructosyltransferase (FBOS) or glucosyltransferase (GBOS) with high concentration of sucrose (3M). MBOS was further modified as iron and sulfate-oligosaccharides. The modified MBOS were stable at high temperatures (up to $140^{\circ}C$) and low pHs (2 to 4). Most highly branched and modified oligosaccharide (0.34%, w/v) effectively inhibited fructose release from sucrose by Streptococcus mutans 6715 mutansucrase. FBOS, GBOS, iron-MBOS inhibited the mutansucrase activities from Streptococcus sobrinus about 46.8%, 49.2% and 43.1%, respectively. Most highly branched and modified oligo- saccharides (0.5%, w/v) effectively inhibited the fonnation of insoluble glucan and adherence of S. mutans or S. sobrinus cell in the presence of sucrose. Modified oligosaccharides affected the growth and acid production of oral pathogens. Cytotoxicity test showed that highly branched and modified oligosaccharides was non-toxic.

• 한국산학기술학회:학술대회논문집
• /
• 한국산학기술학회 2010년도 춘계학술발표논문집 2부
• /
• pp.1221-1223
• /
• 2010

본 논문에서는 한국인 아동의 우식치아로부터 S. mutans를 분리하고, acid stress하에서 분리한 S. mutans의 유전자의 발현의 변화를 분석하고자 하였다. 치아우식증의 주요한 요소로 작용하는 치태형성에 기여하는 glucan 및 fructan 합성에 관여하는 세포내 효소인 glucosyltransferase, glucosyltransferase, glucosyltransferase 및 fructosyltransferase의 발현량의 변화를 확인한 결과, lactic acid를 처리하지 않은 control의 경우보다 16배에서 3배까지 감소한 것을 확인할 수 있었다. Amino acid ABC transporter, adenylate kinase, fructokinase, 40k cell wall protein precursor에서는 모두 유전자의 발현량이 현저히 증가한 것을 볼 수 있었다. 이들 유전자는 acid stress에 관여하는 특이적 유전자로 추정된다.

• Journal of Microbiology and Biotechnology
• /
• 제32권11호
• /
• pp.1373-1381
• /
• 2022

Fructan is a polysaccharide composed of fructose and can be classified into several types, such as inulin, levan, and fructo-oligosaccharides, based on their linkage patterns and degree of polymerization. Owing to its structural and functional diversity, fructan has been used in various fields including prebiotics, foods and beverages, cosmetics, and pharmaceutical applications. With increasing interest in fructans, efficient and straightforward production methods have been explored. Since the 1990s, yeast cells have been employed as producers of recombinant enzymes for enzymatic conversion of fructans including fructosyltransferases derived from various microbes and plants. More recently, yeast cell factories are highlighted as efficient workhorses for fructan production by direct fermentation. In this review, recent advances and strategies for fructan biosynthesis by yeast cell factories are discussed.

• 생명과학회지
• /
• 제25권9호
• /
• pp.1021-1026
• /
• 2015

Invertase (β-D-fructosfuranosidase, EC 3.2.1.26)는 설탕을 포도당과 과당으로 가수분해하는 반응을 촉매한다. 3종류의 invertases [액포(수용성 산), 세포질(수용성 알칼리) 및 세포벽 결합]가 식물에서 연구되어 왔다. 우리는 순차적인 ammonium sulfate 침전, 이온교환크로마토그래피, 흡수크로마토그래피, Green-19 친화크로마토그래피 과정을 통해 완두콩(Pisum sativum L.) 발아체로부터 중성 invertase의 세포막 연결 isoform을 430배 순수 분리하였다. 분리된 세포막과 결합 된insoluble invertase (IN-INV)는 최적 pH는 중성에서 알칼리 사이(pH 6.8-7.5)로 나타났다. 이 효소는 Tris 뿐만 아니라 Hg²⁺ and Cu²⁺와 같은 중금속에 의해 저해되었다. IN-INV 의 K_m과 V_max 값은 각각 12.95 mM과 2.98 U/min으로 측정되었다. IN-INV는 기질로써 과당뿐만 아니라 라피노오스와 반응하기 때문에 진정한 β-fructofuranosidase로 판명되었다. IN-INV의 분자량 20 kDa이었다. 위 결과로 볼 때 GA 영향으로 급속히 자라는 발아체에서 단백질이 분리되었는데 특징적으로 invertase였다.

• Journal of Applied Biological Chemistry
• /
• 제58권3호
• /
• pp.209-218
• /
• 2015

Fructooligosaccharides have been mainly produced by microbial fructosyltransferases (FTase) enzymes. The present work focuses on the optimization of medium composition and cultivation parameters affecting FTase produced by Penicillium aurantiogriseum AUMC 5605 in shake flask cultivation. FTase production was optimized in two steps using DeMeo's fractional factorial design. A 1.46-fold increase in FTase production (105.4 U/mL) was achieved using the optimized culture medium consisting of (g/L): sucrose, 600; yeast extract, 10; $K_2HPO_4$, 5; $MgSO_4{\cdot}7H_2O$, 0.5; $(NH_4)_2SO_4$, 1.0 and KCl, 0.5. The obtained results showed that the maximum FTase enzyme activity was produced at initial cultivation pH values ranging from 6.0-6.5, at agitation speed of 200 rpm and using vegetative fungal cells as inoculum. Moreover, results showed that optimization of medium composition and some cultivation parameters resulted in an increase of about 93.7% in the enzyme activity than the nonoptimized cultivation conditions after 96 h of cultivation. Additionally, maximum production and specific production rates recorded 2340 U/L/h and 102 U/L/h/g cells, respectively.

• International Journal of Oral Biology
• /
• 제44권2호
• /
• pp.31-36
• /
• 2019

Streptococcus mutans is one of the important bacteria that forms dental biofilm and cause dental caries. Virulence genes in S. mutans can be classified into the genes involved in bacterial adhesion, extracellular polysaccharide formation, biofilm formation, sugar uptake and metabolism, acid tolerance, and regulation. The genes involved in bacterial adhesion are gbps (gbpA, gbpB, and gbpC) and spaP. The gbp genes encode glucan-binding protein (GBP) A, GBP B, and GBP C. The spaP gene encodes cell surface antigen, SpaP. The genes involved in extracellular polysaccharide formation are gtfs (gtfB, gtfC, and gtfD) and ftf, which encode glycosyltransferase (GTF) B, GTF C, and GTF D and fructosyltransferase, respectively. The genes involved in biofilm formation are smu630, relA, and comDE. The smu630 gene is important for biofilm formation. The relA and comDE genes contribute to quorumsensing and biofilm formation. The genes involved in sugar uptake and metabolism are eno, ldh, and relA. The eno gene encodes bacterial enolase, which catalyzes the formation of phosphoenolpyruvate. The ldh gene encodes lactic acid dehydrogenase. The relA gene contributes to the regulation of the glucose phosphotransferase system. The genes related to acid tolerance are atpD, aguD, brpA, and relA. The atpD gene encodes $F_1F_0$-ATPase, a proton pump that discharges $H^+$ from within the bacterium to the outside. The aguD gene encodes agmatine deiminase system and produces alkali to overcome acid stress. The genes involved in regulation are vicR, brpA, and relA.

• 한국미생물·생명공학회지
• /
• 제52권2호
• /
• pp.122-134
• /
• 2024

Levan is a functional fructooligosaccharide that belongs to the class of fructans and displays a range of physiological and dietary benefits. Levansucrase, classified as a fructosyltransferase, catalyzes the synthesis of levan fructans through the transfer of d-fructosyl residues from sucrose. Recombinant levansucrase exhibits a significantly high level of expression and activity in E. coli, however, its industrial applicability is limited due to its relatively poor thermal stability. In this study, we generated six single mutants by targeting either the high flexibility regions (242-250) or the 382Glu residue within the -TEAP- motif, and further developed two double mutants featuring concurrent mutations encompassing both regions. We expressed multiple levansucrase mutant proteins in E. coli DH5α using site-directed mutagenesis and measured the enzymatic activity and thermal stability of each mutant by levan formation reactions. The E246Y mutant displayed half-time 5.02-fold and 1.89-fold higher at 55℃ and 56℃, respectively, when compared to the WT, whereas the E382L mutant exhibited a 1.06-fold enhancement in half-time at 55℃. Furthermore, E246Y/E382L exhibited 1.95-fold and 1.86-fold higher thermal stability at 55℃ and 56℃, respectively. These findings suggest the potential for the developed thermally stable levansucrases to be applied in industrial settings, highlighting the effectiveness of our targeted mutagenesis approach.