Hybrid magnetic cross-linked enzyme aggregates of phenylalanine ammonia lyase from Rhodotorula glutinis

Jian dong Cui; Li li Cui; Song ping Zhang; Yu fei Zhang; Zhi guo Su; Guang hui Ma

doi:10.1371/journal.pone.0097221

Hybrid magnetic cross-linked enzyme aggregates of phenylalanine ammonia lyase from Rhodotorula glutinis

PLoS One. 2014 May 13;9(5):e97221. doi: 10.1371/journal.pone.0097221. eCollection 2014.

Authors

Jian dong Cui¹, Li li Cui², Song ping Zhang³, Yu fei Zhang⁴, Zhi guo Su⁴, Guang hui Ma⁴

Affiliations

¹ Research Center for Fermentation Engineering of Hebei, College of Bioscience and Bioengineering, Hebei University of Science and Technology, Shijiazhang, P R China; National Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences, HaiDian district, Beijing, P R China; Key Laboratory of Industry Microbiology, Ministry of Education, Tianjin University of Science and Technology, Tai Da Development Area, Tianjin, P R China.
² Research Center for Fermentation Engineering of Hebei, College of Bioscience and Bioengineering, Hebei University of Science and Technology, Shijiazhang, P R China; National Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences, HaiDian district, Beijing, P R China.
³ National Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences, HaiDian district, Beijing, P R China; Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Tianjin, China.
⁴ National Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences, HaiDian district, Beijing, P R China.

Abstract

Novel hybrid magnetic cross-linked enzyme aggregates of phenylalanine ammonia lyase (HM-PAL-CLEAs) were developed by co-aggregation of enzyme aggregates with magnetite nanoparticles and subsequent crosslinking with glutaraldehyde. The HM-PAL-CLEAs can be easily separated from the reaction mixture by using an external magnetic field. Analysis by scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM) indicated that PAL-CLEAs were inlayed in nanoparticle aggregates. The HM-PAL-CLEAs revealed a broader limit in optimal pH compared to free enzyme and PAL-CLEAs. Although there is no big difference in Km of enzyme in CLEAs and HM-PAL-CLEAs, Vmax of HM-PAL-CLEAs is about 1.75 times higher than that of CLEAs. Compared with free enzyme and PAL-CLEAs, the HM-PAL-CLEAs also exhibited the highest thermal stability, denaturant stability and storage stability. The HM-PAL-CLEAs retained 30% initial activity even after 11 cycles of reuse, whereas PAL-CLEAs retained 35% of its initial activity only after 7 cycles. These results indicated that hybrid magnetic CLEAs technology might be used as a feasible and efficient solution for improving properties of immobilized enzyme in industrial application.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Biotechnology / methods*
Cross-Over Studies
Glutaral / metabolism
Hydrogen-Ion Concentration
Kinetics
Magnetics
Magnetite Nanoparticles / chemistry*
Microscopy, Confocal
Microscopy, Electron, Scanning
Phenylalanine Ammonia-Lyase / metabolism*
Protein Aggregates / physiology*
Rhodotorula / enzymology*
Rhodotorula / physiology

Substances

Magnetite Nanoparticles
Protein Aggregates
Phenylalanine Ammonia-Lyase
Glutaral

Grants and funding

The project was partially supported by the National Natural Science Foundation of China (project no. 21072041, 21106164, 21376249). Dr. J.D. Cui also thanks supports from Open Funding Project of the National Key Laboratory of Biochemical Engineering (no. KF2010-12) and the Foundation (no. 2012IM004) of Key Laboratory of Industry Microbiology, Ministry of Education (Tianjin Key Laboratory of Industrial Microbiology), (Tianjin University of Science & Technology), People’s Republic of China, the Natural Science Foundation of Hebei Province, China (project no. B2014208054), and the Foundation of Hebei University of Science and Technology (project no. SW10). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.