TY - JOUR
T1 - Nano-fibrillated cellulose-based scaffolds for enzyme (co)-immobilization
T2 - Application to natural product glycosylation by Leloir glycosyltransferases
AU - Liu, Hui
AU - Štiglic, Andrea Dobaj
AU - Mohan, Tamilselvan
AU - Kargl, Rupert
AU - Kleinschek, Karin Stana
AU - Nidetzky, Bernd
N1 - Publisher Copyright:
© 2022 The Authors
PY - 2022/12/1
Y1 - 2022/12/1
N2 - Polysaccharide-based scaffolds are promising carriers for enzyme immobilization. Here, we demonstrate a porous scaffold prepared by direct-ink-writing 3D printing of an ink consisting of nanofibrillated cellulose, carboxymethyl cellulose and citric acid for immobilization application. Negative surface charge introduced by the components made the scaffold amenable for an affinity-like immobilization via the cationic protein module Zbasic2. Zbasic2 fusions of two sugar nucleotide-dependent glycosyltransferases (C-glycosyltransferase, Z-CGT; sucrose synthase, Z-SuSy) were immobilized individually, or co-immobilized, and applied to synthesize the natural C-glycoside nothofagin. The cascade reaction involved β-C-glycosylation of phloretin (10 mM, ~90 % conversion) from UDP-glucose, provided from sucrose and catalytic amounts of UDP (1.0 mM). Enzymes were co-immobilized at ~65 mg protein/g carrier to receive activities of 9.5 U/g (Z-CGT) and 4.5 U/g (Z-SuSy) in 22–33 % yield (protein) and an effectiveness of 23 % (Z-CGT) and 13 % (Z-SuSy). The scaffold-bound enzymes were recyclable for 5 consecutive reactions.
AB - Polysaccharide-based scaffolds are promising carriers for enzyme immobilization. Here, we demonstrate a porous scaffold prepared by direct-ink-writing 3D printing of an ink consisting of nanofibrillated cellulose, carboxymethyl cellulose and citric acid for immobilization application. Negative surface charge introduced by the components made the scaffold amenable for an affinity-like immobilization via the cationic protein module Zbasic2. Zbasic2 fusions of two sugar nucleotide-dependent glycosyltransferases (C-glycosyltransferase, Z-CGT; sucrose synthase, Z-SuSy) were immobilized individually, or co-immobilized, and applied to synthesize the natural C-glycoside nothofagin. The cascade reaction involved β-C-glycosylation of phloretin (10 mM, ~90 % conversion) from UDP-glucose, provided from sucrose and catalytic amounts of UDP (1.0 mM). Enzymes were co-immobilized at ~65 mg protein/g carrier to receive activities of 9.5 U/g (Z-CGT) and 4.5 U/g (Z-SuSy) in 22–33 % yield (protein) and an effectiveness of 23 % (Z-CGT) and 13 % (Z-SuSy). The scaffold-bound enzymes were recyclable for 5 consecutive reactions.
KW - 3D printing
KW - Carboxymethyl cellulose
KW - Enzyme (co)immobilization
KW - Leloir glycosyltransferase
KW - Nano-fibrillated cellulose
KW - Porous scaffolds
UR - http://www.scopus.com/inward/record.url?scp=85138594844&partnerID=8YFLogxK
U2 - 10.1016/j.ijbiomac.2022.09.160
DO - 10.1016/j.ijbiomac.2022.09.160
M3 - Article
C2 - 36165869
SN - 0141-8130
VL - 222
SP - 217
EP - 227
JO - International Journal of Biological Macromolecules
JF - International Journal of Biological Macromolecules
IS - Pt A
ER -