KSBB Journal

The Korean Society for Biotechnology and Bioengineering (한국생물공학회)
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Isolation and Purification of Methyl Mercaptan Oxidase from Thiobacillus thiooxidans for Detection of Mercaptasn

머캅탄류 검출을 위한 Thiobacillus thiooxidans가 생산하는 메칠머캅탄 산화효소의 분리 및 정제

  • 김상준 (한국과학기술원 화학공학과 생물환경공학연구실) ;
  • 신현재 (생명공학연구소 당생물학 연구실) ;
  • 이대실 (생명공학연구소 당생물학 연구실) ;
  • 양지원 (한국과학기술원 화학공학과 생물환경공학연구실)
  • Published : 2000.04.01
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Abstract

Methyl mercaptan oxidase was isolated and purified from Thiobacillus thiooxidans KCTC2505 for the detection of mercaptans. T The produre of purification involved DEAE-Sephacel and Superose 12 column chromatographies with recovery yields of 40 a and 6.3%, and specific activity of 19.7 and 80.1 units/mg-protein, respectively. The molecular weight of purified methyl m mercaptan oxidase was determined to be 68.1 kDa by SDS-PAGE. The extract from DEAE-Sephacel column chromatography h had a high activity in oxidizing methyl mercaptan to produce formaldehyde which can be easily detected by purpald-coloring m method. Optimum temperature for activity was observed at $43^{\circ}C$. This enzyme was activated by $NH_4CI and (NH_4)_2S0_4$, and | inhibited by KCI and NaC!.

Thiobacillus thiooxidans KCTC2505에서 메칠머캅탄(MM) 가스를 기질로 메칠머캅탄 산화효소(MMO)를 유도할 수 있었으며 DEAE-Sephacel과 Superose 12 컬럼 크로마토그래피를 이용하여 정제하였다. 이때 얻어진 각 효소의 비활성은 각각 19.7과 80.1 units/mg-protein이었으며 효소의 최적 온도는 $43^{\circ}C$였다. 상기 정제된 효소액은 SDS-PAGE상에서 68.1 kDa의 분자량을 가진다. 이 효소는 암모늄염의 존재하에서는 활성이 증가하였으나 KCl 존재하에서는 감소하였으며 NaCl에 대해서는 거의 영향을 받지 않는 특성을 가지고 있다. 에탄올, 메탄올, 글리세린의 첨가에 대해 효소활성은 부분적으로 불활성화되었으며 아세톤의 경우에는 완전히 저해되었다. Purpald 발색법을 이용한 흡광도의 변화는 구강내 존재하는 MM 가스로부터 생성될 수 있는 수 십 nmol의 범위의 포름알데하이드에 대해 눈으로 인지가능한 발색시스템에 사용할 수 있음을 확인하였다.

Keywords

References

  1. J. Peridontol v.63 Measurement of oral malodors: current methods and future prospects Rosenberg, M.;Christopher A.G. McCulloch
  2. AJVR v.59 Otal malodor measurements on a tooth surface of dogs with gingivitis Hennet;Philippe R.;Beatrice Delille;Jean Louis Davot
  3. J. Periodontol v.69 The effect of a 1-stage full-mouth disinfection on oral malodor and microbial colonization of the rongue in peridontitis patients. A pilot study Quirynen, M.;C. Mongaridini;D. van Steenberghe
  4. US patent 4119089 Method of predicting and determining ovulation by monitoring the concentration of volatile sulfur-containing compounds present in mouth air Preti, G.;G.R. Huggins;J. Tonzetich
  5. J. Gen. Microb. v.138 Purification and properties of methyl mercaptan oxidase from Thiobacillus thiparus TK-m Gould, W.D.;T. Kanagawa
  6. J. Gen. Microb. v.127 Dimethyl Sulphoxide and dimethyl sulphide as a carbon, sulphur and energy source for growth of Hyphomicrobium S Bont, J.A.M.De;J.P. Dijken;W. Harder
  7. J. Gen. Microb. v.133 Methyl mercaptan oxidase, a key enzyme in the metabolism of methylated sulphur compounds by Hyphomicrobium EG Suylen, G.M.H.;P.J. Large;J.P. Van Dijken;J.G. Kuenen
  8. Analytical Biochemistry v.234 A rapid formaldehyde assay using purpald reagent: Application under periodation congition Quesenberry, M.S.;Y.C. Lee
  9. Analytical Biochemistry v.267 Quantification of bacterial lipopolysaccharides by the purpald assay: measuring formaldehyde generated from 2-keto-3- deoxyoctonate and heptose at the inner core by periodate oxidation Lee, C.H.;C.M. Tsai
  10. Russian J. Applied Chemistry v.72 Kinetics and pathways of oxidation of o-phenylenediamine, 3,3-dimethoxybenzidine, and 3,3,5,5-tetramethylbenzidine with hydrogen peroxide, catalyzed with horse radish peroxidase immobilized on various supports Muginova, S.V.;I.A. Veselova;T.N. Shekhovtsova
  11. J. Analytical Chemistry v.54 Use of paper as a support for immobilized peroxidase in a test method for the deterination of mercury(II) traces Mugunova, S.V.;N.A. Akovbyan;T.N. Shekhovtsova
  12. J. Fermentation and Bioengineering v.6 Characteristics of hydrogen sulfide removal by Thiobacillus thiooxidans KS1 isolated from a crrier-packed biological deodorization system Shinabe, K.;S. Oketani;T. Ochi;M. Matsumura
  13. Ph.D. Dissertation Isolation and deodorizing characteristics of sulfur oxidizing bacteria and ammonia oxdizing bacteria for the removal of sulfur containing malodorous compounds and ammonia Lee, U.Y.
  14. Analytical Biochemistry v.72 A rapid and sensitive mehod for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding Bradford, M.M.
  15. Process Biochemistry v.34 Bulk and surface characterization of crystalline and plastic sulphur oxidized by two Thiobacillus species Briand, L.E.;R.D. Bonetto;J.L. Ladaga;E. Donati
  16. J. Korea Ind. & Chemistry v.5 A study on the preparation of polyurethane diagnotic membrane for urine glucose test Kwon, S.K.