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최태호,조남석,민두식,오가쇼지,구창덕,안드레 레오노비치 한국목재공학회 2000 목재공학 Vol.28 No.1
Culture maturity assessment can be used to control fruiting body flush timing. Culture maturity of sawdust-based substrate was evaluated by using oak mushroom, [Lentinula edodes (Berk.) Pegler]. The influence of substrate water potential (ψ) on the growth and fruiting of three genotypes of L. edodes was also investigated. Glucosamine content revealed a peak at the fruiting body senescent stage. Glucosamine increased steadily to the sporophore senescent stage, and sharply declined at crop treatment. Lipid phosphate and ergosterol contents peaked at pinning and button break stages, respectively. Therefore lipid phosphate and ergosterol contents would be considered as the convenient measurement for judging culture maturity and fruiting potentials. The substrate pH values before inoculation and on the fruiting stage were varied from 6.3 to 4.0. This pH changes were detected as changes in color from bluish purple to yellow by direct bromphenol blue(BPB) spraying, and shown a good correlation with fruit body yield of the 1st flush. Concerning water potential of the cultures, a slight reduction of water potential, -0.5MPa, stimulated mycelial and colony growths on liquid, agar and sawdust-based substrates. The water potential of well-colonized matured substrate was -0.7MPa and -4.0MPa, before and after the fruiting, respectively. Excellent water providing capacity (higher ψ) is expected to well-matured cultures with a high density of mycelial colonization. Also, the substrate water potential significantly affected by the interaction between genotypes and spawn run time.
2,5-Xylidine 을 이용한 목재부후균으로부터 Laccase 효소의 유도
조남석,김영신,방명혁,최윤정,남장현,안드레레오노비취 한국목재공학회 1998 목재공학 Vol.26 No.3
Some white-rot fungi, screened at the Laboratory of Forest Products Microbiological Chemistry, Chungbuk National University were cultured and added the inducer of lactase enzyme, 2,5-xylidine. The fungi named by CB-13, CB-20, CB-99, CB-100 and CB-123 strains showed positive results in the decolorization of aromatic compounds, carminic acid and Rhemazol brilliant blue R. Concerned to the inducing effect of 2,5-xylidine on lactase activity, CB-20, CB-100 and CB-123 strains showed very high activity by addition of 2,5-xylidine, whilst CB-13, CB-99 and CB-124 strains produced relatively high lactase enzymes, regardless of inducer addition. There were no any lactase activities on CB-25, CB-64 and CB-139, even in addition of inducer. It is confirmed that some screened fungi have decolorizing ability on aromatic compounds, carminic acid and Rhemazol brilliant blue R. Also, the addition of inducer, 2,5-xylidine, has increased the activity of lactase enzyme which is secreted from some white-rot fungi.
Fungal laccases from basidiomycetes and their inducibility
Andrzej Leonowicz,A. Wilkolazka,J. Rogalski,Dong-Hoon Kim,Nam-Seok Cho 한국버섯학회 2004 한국버섯학회지 Vol.2 No.3
Laccases are multicopper-containing enzymes which catalyze the oxidation of phenolic and nonphenolic compounds with the concomitant reduction of molecular oxygen. They often occur as isoenzymes, either constitutive or inducible, that oligomerize to multilateral complexes, what allow for penetration to the woody cell wall structure. White rot basidiomycete fungi may produce a number of laccase isoenzymes, some constitutively and others after induction. Fungal laccase is commonly induced by many ions, such as Cu2+, Cd2+ Ca2+, Li+, Mn2+, Ag+, Hg2+, Mn and Fe3+, phenolic compounds, some organic compounds, such as ethanol, isopropanol, cAMP, caffeine, p-anisidine, viscosinamide and paraquat, and nitrogens and even heat shock. A combination of Cu and pHB (p-hydroxybenzoic acid) made it possible to extend the inducible laccase activities over 30-fold. But the most effective inducer of laccase in the basidiomycete and other higher fungi is 2,5-xylidine, over 160-fold stimulation of laccase activity. The laccases are frequently encoded by gene families, as e.g. in Pycnoporus cinnabarinus, from which the lcc3-1 or the allelic form lac1 and lac3-2 have been cloned and sequenced. In the case of inducible forms the post-inductional laccase formation depends upon the synthesis of mRNA and the induction is due to the synthesis of a new protein.
Fungal laccases from basidiomycetes and their inducibility
안드레 레오노비치,엥 빌코오즈카,제이 로갈스키,김동훈,조남석,Leonowicz, Andrzej,Wilkolazka, A.,Rogalski, J.,Kim, Dong-Hoon,Cho, Nam-Seok The Korean Society of Mushroom Science 2004 한국버섯학회지 Vol.2 No.3
Laccase는 여러개의 Cu를 포함하는 효소로서 분자상 산소를 환원시키면서 패놀성 및 비페놀성화합물의 산화를 촉매하는 작용을 한다. 이들 효소들은 미생물 고유의 혹은 유도상의 동위효소의 형태로 목질화된 세포벽을 침투하게 된다. 백색부후균은 많은 종류의 고유의 혹은 유도상의 동위효소를 생산한다. 이들 균체 laccase효소들은 통상 $Cu^{2+}$, $Cd^{2+}$ $Ca^{2+}$, $Li^+$, $Mn^{2+}$, $Ag^+$, $Hg^{2+}$, Mn 및 $Fe^{3+}$이온과 같은 금속이온들, 페놀성 화합물, ethanol, isopropanol, cAMP, caffeine, p-anisidine, viscosinamide 및 paraquat 등과 같은 유기화합물, 질소 및 열충격 등에 의하여 유도될 수 있다. Cu 및 pHB (p-hydroxybenzoic acid)의 조합으로 laccase 활성을 30배이상 유도시킬 수 있었다. 여러 가지 inducer 가운데, 2,5-xylidine이 담자균 및 기타 다른 고등균류로부터 160배이상의 가장 효과적인 laccase의 유도효과를 나타냈다. 한편 laccase효소는 Pycnoporus cinnabarinus의 gene family로부터 자주 code로 표시되었는데, lcc3-1 혹은 lac1 및 lac3-2의 페어 gene으로 클론 및 시퀀싱되었다. 유도형 laccase의 경우 mRNA의 합성에 의존하여 laccase가 생성되며, 이러한 유도효과는 결국 새로운 단백질의 합성으로부터 기인된다. Laccases are multicopper-containing enzymes which catalyze the oxidation of phenolic and nonphenolic compounds with the concomitant reduction of molecular oxygen. They often occur as isoenzymes, either constitutive or inducible, that oligomerize to multilateral complexes, what allow for penetration to the woody cell wall structure. White rot basidiomycete fungi may produce a number of laccase isoenzymes, some constitutively and others after induction. Fungal laccase is commonly induced by many ions, such as $Cu^{2+}$, $Cd^{2+}$ $Ca^{2+}$, $Li^+$, $Mn^{2+}$, $Ag^+$, $Hg^{2+}$, Mn and $Fe^{3+}$, phenolic compounds, some organic compounds, such as ethanol, isopropanol, cAMP, caffeine, p-anisidine, viscosinamide and paraquat, and nitrogens and even heat shock. A combination of Cu and pHB (p-hydroxybenzoic acid) made it possible to extend the inducible laccase activities over 30-fold. But the most effective inducer of laccase in the basidiomycete and other higher fungi is 2,5-xylidine, over 160-fold stimulation of laccase activity. The laccases are frequently encoded by gene families, as e.g. in Pycnoporus cinnabarinus, from which the lcc3-1 or the allelic form lac1 and lac3-2 have been cloned and sequenced. In the case of inducible forms the post-inductional laccase formation depends upon the synthesis of mRNA and the induction is due to the synthesis of a new protein.
조남석,안드레레오노비취,리리아나지안프레다,죠오지로갈스키,마그다야색,욜란타루테렉,마리아바실레브스카,엘리지베타말라책,안드레다비도비취,모니카핑크브츠,그리지나기날스카,마그다스타스책 한국목재공학회 1997 목재공학 Vol.25 No.3
목재부후균으로부터 락케이스 효소의 생산 및 유도를 위하여 여러 가지 유도약품(inducer)을 사용하였다. 이들 가운데 ferulic acid, pentachlorophenol 및 2,5-xylidine이 매우 높은 락케이스 활성을 나타나게 하였으며 거의 동일한 유도효과를 보여주었다. 이들 약품 이외에도 sinapic acid syringic acid 및 coffeic acid 등도 높은 락케이스 활성을 주었는데, 산의 형태가 알데히드류보다도 높은 유도효과를 나타냈다. 그리고 실험한 48개 균주가운데 38개 균주가 락케이스를 생산하였으며, 이 가운데 32균주가 ferulic acid에 의해 강한 효소유도 활성을 보였다. 이러한 결과는 지금까지 락케이스 효소의 검출이 어려웠던 Abortiporus biennis 및 Gleophyllum odoratum에서도 높은 락케이스 효소의 유도를 가능하게 하였다. 아울러 가장 높은 효소활성을 나타낸 균주로서는 Cerrena unicolor 였으며, 그 락케이스 효소활성이 무처리 및 inducer 첨가시 각각 40,000 및 60,000 nkat/l 정도 였다.
Purification of the Candida utilis Extracellular Invertase using Affinity Chromatography
G.Ginalska,조남석,A.Belcarz,J.Lobarzewski,A.Leonowicz 한국목재공학회 2002 목재공학 Vol.30 No.3
The extracellular invertase (EC 3.2.1.26) (Candida utilis) preparation was obtained from the liquid medium after desalting and freeze drying. This prepared enzyme was used for the comparative purification on 4 activated matrices by liquid column affinity chromatography method. In this method there were used controlled porous glass (CPG) silanized covalently activated by keratin, silanized silica gel and silica gel covalently covered by keratin. It was found that the invertase purification process was better using both CPG matrices (silanized CPG and keratin activated CPG) than these with two silica gel supports. Also the elution coefficient of the invertase from the two CPG columns was about 93 to 94%. Two silica gel supports found to be superior in terms of purification efficiency. The invertase purification process was confirmed by PAGE electrophoresis.