공유결합으로 다공성 막에 고정화된 PLD에 의한 포스퍼티딕산 생산

박진원; Jin-Won Park

Clean Technology, Vol.21, No.4, 224-228, December, 2015

DOI10.7464/ksct.2015.21.4.224 Export Citation

공유결합으로 다공성 막에 고정화된 PLD에 의한 포스퍼티딕산 생산

Phosphatidic Acid Production by PLD Covalently Immobilized on Porous Membrane

박진원^†

서울과학기술대학교 화공생명공학과, 01811 서울특별시 노원구 공릉로 232

Jin-Won Park^†

Department of Chemical and Biomolecular Engineering, College of Energy and Biotechnology, Seoul National University of Science and Technology, Gongreung-ro 232, Nowon-gu, Seoul 01811, Korea

E-mail:

초록

포스포라이페이즈디를 공유결합을 통해서 초미세다공성막에 고정화하였다. 고정화는 폴리에틸렌이민, 글루타알데하이드, 포스포라이페이즈디를 순차적으로 처리함으로써 수행되었다. X선 광전자 분광기를 이용하여 고정화가 확인되었다. 포스퍼티딜콜린이 분산된 버퍼용액의 pH값을 시간에 따라 모니터링하여 고정화된 경우와 그렇지 않은 경우에 대해 촉매활성을 산출하였다. 속도상수는 폴리스타이렌나노입자에 고정화된 포스포라이페이즈디에서는 0.64 s-1, 다공성 셀룰로스아세테이트막에 고정화된 포스포라이페이즈디에서는 0.52 s-1, 그리고 고정화되지 않은 포스포라이페이즈디에서는 0.75 s-1의 결과가 도출되었다. 재사용에 대한 연구가 10차례까지 수행되었으며, 초기 사용시의 활성대비로 95%가 유지되었다. 열과 저장성에 대한 안정성도 고찰되었으며, 다공성막에 고정화된 포스포라이페이즈디의 경우에 활성손실이 가장 적은 것으로 관찰되었다. 이 연구결과들로부터, 포스퍼티딕산의 생산용 포스포라이페이즈디의 고정화에 대한 지지체로 다공성 막을 사용할 수 있음을 알 수 있다.

Phospholipase D (PLD) was immobilized on a submicro-porous membrane through covalent immobilization. The immobilization was conducted on the porous membrane surface with the treatment of polyethyleneimine, glutaraldehyde, and the anhydrase, in sequence. The immobilization was confirmed using X-ray photon spectrometer. The pH values of phosphatidylcholine (PC) dispersion solution with buffer were monitored with respect to time to calculate the catalytic activities of PC for free and immobilized PLD. The catalytic rate constant values for free PLD, immobilized PLD on polystyrene nanoparticles, and immobilized PLD on a porous cellulose acetate membrane were 0.75, 0.64, and 0.52 s-1, respectively. Reusability was studied up to 10 cycles of PC hydrolysis. The activity for the PLD immobilized on the membrane was kept to 95% after 10 cycles, and comparable to the PLD on the nanoparticles. The stabilities for heat and storage were also investigated for the three cases. The results suggested that the PLD immobilized on the membrane had the least loss rate of the activity compared to the others. From these studies, the porous membrane was feasible as a carrier for the PLD immobilization in the production of phosphatidic acid.

Keywords:Phospholipase D immobilization;Porous membrane;Phosphatidyl choline;Hydrolysis;Phosphatidic acid

[References]

Delon C, Manifava M, Wood E, Thompson D, Krugmann S, Pyne S, Ktistakis NT, J. Biol. Chem., 279, 44763 (2004)
Raghu P, Biochim. Biophys. Acta, 1821, 1154 (2012)
Weigert R, Silletta MG, Spano S, Turacchio G, Cericola C, Colanzi A, Senatore S, Mancini R, Polishchuk EV, Salmona M, Facchiano F, Burger KN, Mironov A, Luini A, Corda D, Nature, 402, 429 (1999)
Blackwood RA, Smolen JE, Transue A, Hessler RJ, Harsh DM, Brower RC, French S, Am. J. Physiol., 272, C1279 (1997)
Shemesh T, Luini A, Malhotra V, Burger KN, Kozlov MM, Biophys. J., 85, 3813 (2003)
Kooijman EE, Chupin V, de Kruijff B, Burger KN, Traffic, 4, 162 (2003)
Athenstaedt K, Daum G, Eur. J. Biochem., 266, 1 (1999)
Brindley DN, Waggoner DW, Chem. Phys. Lipids, 80, 45 (1996)
Park JW, J. Membr. Sci., 246, 399 (2013)
Gu HA, Kim MJ, Kim HS, Ha JH, Yu ER, Park SN, Appl. Chem. Eng., 26(2), 165 (2015)
Park SN, Jo NR, Jeon SH, J. Ind. Eng. Chem., 20(4), 1481 (2014)
Schroeder R, London E, Brown D, Proc. Natl. Acad. Sci. USA, 91, 12130 (1994)
Grun A, Limpacher R, Ber. Der. Deutschen Chem. Geser., 59b, 1345 (1926)
Liscovitch M, Czarny M, Fiucci G, Tang X, Biochem. J., 345, 401 (2000)
Chong PLG, Sugar IP, Chem. Phys. Lipids, 116, 153 (2002)
Vinoba M, Bhagiyalakshmi M, Jeong SK, Yoon Y, Nam SC, Colloids Surf. B: Biointerfaces, 90, 91 (2012)
Lee JM, Biochemical Engineering, Prentice Hall, Englewood Cliffs, NJ, USA, 1992, 21-31.
Martinezdiez L, Vazquezgonzalez MI, AIChE J., 42(7), 1844 (1996)

[Referenced By]

[1] Delon C, Manifava M, Wood E, Thompson D, Krugmann S, Pyne S, Ktistakis NT, J. Biol. Chem., 279, 44763 (2004)

[2] Raghu P, Biochim. Biophys. Acta, 1821, 1154 (2012)

[3] Weigert R, Silletta MG, Spano S, Turacchio G, Cericola C, Colanzi A, Senatore S, Mancini R, Polishchuk EV, Salmona M, Facchiano F, Burger KN, Mironov A, Luini A, Corda D, Nature, 402, 429 (1999)

[4] Blackwood RA, Smolen JE, Transue A, Hessler RJ, Harsh DM, Brower RC, French S, Am. J. Physiol., 272, C1279 (1997)

[5] Shemesh T, Luini A, Malhotra V, Burger KN, Kozlov MM, Biophys. J., 85, 3813 (2003)

[6] Kooijman EE, Chupin V, de Kruijff B, Burger KN, Traffic, 4, 162 (2003)

[7] Athenstaedt K, Daum G, Eur. J. Biochem., 266, 1 (1999)

[8] Brindley DN, Waggoner DW, Chem. Phys. Lipids, 80, 45 (1996)

[9] Park JW, J. Membr. Sci., 246, 399 (2013)

[10] Gu HA, Kim MJ, Kim HS, Ha JH, Yu ER, Park SN, Appl. Chem. Eng., 26(2), 165 (2015)

[11] Park SN, Jo NR, Jeon SH, J. Ind. Eng. Chem., 20(4), 1481 (2014)

[12] Schroeder R, London E, Brown D, Proc. Natl. Acad. Sci. USA, 91, 12130 (1994)

[13] Grun A, Limpacher R, Ber. Der. Deutschen Chem. Geser., 59b, 1345 (1926)

[14] Liscovitch M, Czarny M, Fiucci G, Tang X, Biochem. J., 345, 401 (2000)

[15] Chong PLG, Sugar IP, Chem. Phys. Lipids, 116, 153 (2002)

[16] Vinoba M, Bhagiyalakshmi M, Jeong SK, Yoon Y, Nam SC, Colloids Surf. B: Biointerfaces, 90, 91 (2012)

[17] Lee JM, Biochemical Engineering, Prentice Hall, Englewood Cliffs, NJ, USA, 1992, 21-31.

[18] Martinezdiez L, Vazquezgonzalez MI, AIChE J., 42(7), 1844 (1996)