Surface-functionalized hyperbranched poly(amido acid) magnetic nanocarriers for covalent immobilization of a bacterial γ-glutamyltranspeptidase.
In this study, we synthesized water-soluble hyperbranched poly(amido acid)s (HBPAAs) featuring multiple terminal CO2H units and internal tertiary amino and amido moieties and then used them in conjunction with an in situ Fe2+/Fe3+ co-precipitation process to prepare organic/magnetic nanocarriers comprising uniformly small magnetic iron oxide nanoparticles (NP) incorporated within the globular HBPAAs. Transmission electron microscopy revealed that the HBPAA-γ-Fe2O3 NPs had dimensions of 6-11 nm, significantly smaller than those of the pristine γ-Fe2O3 (20-30 nm). Subsequently, we covalently immobilized a bacterial γ-glutamyltranspeptidase (BlGGT) upon the HBPAA-γ-Fe2O3 nanocarriers through the formation of amide linkages in the presence of a coupling agent. Magnetization curves of the HBPAA-γ-Fe2O3/BlGGT composites measured at 300 K suggested superparamagnetic characteristics, with a saturation magnetization of 52 emu g⁻¹. The loading capacity of BlGGT on the HBPAA-γ-Fe2O3 nanocarriers was 16 mg g⁻¹ support; this sample provided a 48% recovery of the initial activity. The immobilized enzyme could be recycled 10 times with 32% retention of the initial activity; it had stability comparable with that of the free enzyme during a storage period of 63 days. The covalent immobilization and stability of the enzyme and the magnetization provided by the HBPAA-γ-Fe2O3 NPs suggests that this approach could be an economical means of depositing bioactive enzymes upon nanocarriers for BlGGT-mediated bio-catalysis.
Department of Applied Chemistry, National Chiayi University, Chiayi 60004, Taiwan. tyjuang@mail.ncyu.edu.tw.
Tzong-Yuan
TY
Juang
Department of Applied Chemistry, National Chiayi University, Chiayi 60004, Taiwan.
Shao-Ju
SJ
Kan
Department of Applied Chemistry, National Chiayi University, Chiayi 60004, Taiwan.
Yi-Yu
YY
Chen
Department of Applied Chemistry, National Chiayi University, Chiayi 60004, Taiwan.
Yi-Lin
YL
Tsai
Institute of Molecular Biology, Academia Sinica, Nankang District, Taipei 11529, Taiwan.
Min-Guan
MG
Lin
Department of Applied Chemistry, National Chiayi University, Chiayi 60004, Taiwan. llin@mail.ncyu.edu.tw.
Long-Liu
LL
Lin
D001426:Bacterial Proteins / Q000737:chemistry*; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000737:chemistry*; D017743:Equipment Reuse; D004926:Escherichia coli / Q000737:chemistry / Q000201:enzymology / Q000235:genetics; D005290:Ferric Compounds / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D058185:Magnetite Nanoparticles / Q000737:chemistry*; D009757:Nylons / Q000737:chemistry*; D010316:Particle Size; D011994:Recombinant Proteins / Q000737:chemistry; D013696:Temperature; D005723:gamma-Glutamyltransferase / Q000737:chemistry*
D001426:Bacterial Proteins; D004800:Enzymes, Immobilized; D005290:Ferric Compounds; D058185:Magnetite Nanoparticles; D009757:Nylons; D011994:Recombinant Proteins; C000499:ferric oxide; D005723:gamma-Glutamyltransferase