Enzymatically active fibrils generated by the self-assembly of the ApoA-I fibrillogenic domain functionalized with a catalytic moiety.
Enzymatically active fibrils were produced by self-assembly of a bifunctional chimeric protein, made up of a fibrillogenic and a catalytic moiety. For this purpose, the fibrillogenic domain of Apolipoprotein A-I (ApoA-I), a 93-residue polypeptide named [1-93]ApoA-I, was functionalized with the enzyme glutathione S-transferase (GST). The fusion protein GST-[1-93]ApoA-I was expressed, isolated to homogeneity and characterized. In the soluble form, GST-[1-93]ApoA-I was found to be fully active as a GST enzyme, and to have high propensity to self-aggregate. Upon incubation for 3 weeks at pH 6.4, insoluble aggregates were generated. Analyzed by AFM, they were found to contain fibrillar structures often organized into large fiber networks. Fibrils were loaded on the membrane of a microfiltration unit and tested for enzymatic activity by filtering the substrate through the fibrillar network. Fibrils were shown to be catalytically active, stable over time and reusable, as no loss of activity was detected when fibrils were repeatedly tested. Our findings suggest that catalytically active fibrils may be of interest for biocatalytic applications in nanobiotechnology.
Guglielmi
Fulvio
F
Department of Structural and Functional Biology, School of Biotechnological Sciences, University of Naples Federico II, Naples 80126, Italy.
Monti
Daria M
DM
Arciello
Angela
A
Torrassa
Silvia
S
Cozzolino
Flora
F
Pucci
Piero
P
Relini
Annalisa
A
Piccoli
Renata
R
D000682:Amyloid / Q000737:chemistry* / Q000378:metabolism*; D016632:Apolipoprotein A-I / Q000737:chemistry* / Q000378:metabolism*; D001665:Binding Sites; D045744:Cell Line, Tumor; D002942:Circular Dichroism; D019281:Dimerization; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D018625:Microscopy, Atomic Force; D046912:Multiprotein Complexes / Q000737:chemistry / Q000378:metabolism; D036103:Nanotechnology; D011485:Protein Binding
D000682:Amyloid; D016632:Apolipoprotein A-I; D046912:Multiprotein Complexes; D005982:Glutathione Transferase