Immobilization of cross-linked phenylalanine ammonia lyase aggregates in microporous silica gel

Jian Dong Cui; Lian Lian Li; Hong Jie Bian

doi:10.1371/journal.pone.0080581

Immobilization of cross-linked phenylalanine ammonia lyase aggregates in microporous silica gel

PLoS One. 2013 Nov 15;8(11):e80581. doi: 10.1371/journal.pone.0080581. eCollection 2013.

Authors

Jian Dong Cui¹, Lian Lian Li, Hong Jie Bian

Affiliation

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.

Abstract

A separable and highly-stable enzyme system was developed by adsorption of phenylalanine ammonia lyase (PAL) from Rhodotorula glutinis in amino-functionalized macroporous silica gel and subsequent enzyme crosslinking. This resulted in the formation of cross-linked enzyme aggregates (PAL-CLEAs) into macroporous silica gel (MSG-CLEAs). The effect of adsorptive conditions, type of aggregating agent, its concentration as well as that of cross-linking agent was studied. MSG-CLEAs production was most effective using ammonium sulfate (40%-saturation), followed by cross-linking for 1 h with 1.5% (v/v) glutaraldehyde. The resulting MSG-CLEAs extended the optimal temperature and pH range compared to free PAL and PAL-CLEAs. Moreover, MSG-CLEAs exhibited the excellent stability of the enzyme against various deactivating conditions such as temperature and denaturants, and showed higher storage stability compared to the free PAL and the conventional PAL-CLEAs. Such as, after 6 h incubation at 60°C, the MSG-CLEAs still retained more than 47% of the initial activity whereas PAL-CLEAs only retained 7% of the initial activity. Especially, the MSG-CLEAs exhibited good reusability due to its suitable size and active properties. These results indicated that PAL-CLEAs on MSG 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

Adsorption
Cross-Linking Reagents / chemistry
Enzyme Activation
Enzymes, Immobilized / chemistry*
Hydrogen-Ion Concentration
Kinetics
Phenylalanine Ammonia-Lyase / chemistry*
Porosity
Rhodotorula / enzymology
Silica Gel / chemistry*
Spectroscopy, Fourier Transform Infrared
Surface Properties
Temperature
Thermodynamics

Substances

Cross-Linking Reagents
Enzymes, Immobilized
Silica Gel
Phenylalanine Ammonia-Lyase

Grants and funding

The authors are grateful for the financial support provided by the National Natural Science Foundation of China (NSFC, Project No. 21072041), Open Funding Project of the National Key Laboratory of Biochemical Engineering (NO. KF2010-12) and the Foundation (NO. 2012IM004) of Tianjin Key Laboratory of Industrial Microbiology (Tianjin University of Science and Technology), P. R. China, and Foundation of Hebei University of Science and technology for Distinguished Young Scientists. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.