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|Poly-3-Hydroxybutyrate Functionalization with BioF-Tagged Recombinant Proteins
|Bello Gil, Daniel
Prieto, M. Auxiliadora
Sanz, Jesús M.
|American Society for Microbiology
|Polyhydroxyalkanoates (PHAs) are biodegradable polyesters that accumulate in the cytoplasm of certain bacteria. One promising biotechnological application utilizes these biopolymers as supports for protein immobilization. Here, the PHA-binding domain of the Pseudomonas putida KT2440 PhaF phasin (BioF polypeptide) was investigated as an affinity tag for the in vitro functionalization of poly-3-hydroxybutyrate (PHB) particles with recombinant proteins, namely, full-length PhaF and two fusion proteins tagged to BioF (BioF-C-LytA and BioF-beta-galactosidase, containing the choline-binding module C-LytA and the beta-galactosidase enzyme, respectively). The protein-biopolyester interaction was strong and stable at a wide range of pHs and temperatures, and the bound protein was highly protected from self-degradation, while the binding strength could be modulated by coating with amphiphilic compounds. Finally, BioF-beta-galactosidase displayed very stable enzymatic activity after several continuous activity-plus-washing cycles when immobilized in a minibioreactor. Our results demonstrate the potentialities of PHA and the BioF tag for the construction of novel bioactive materials. IMPORTANCE Our results confirm the biotechnological potential of the BioF affinity tag as a versatile tool for functionalizing PHA supports with recombinant proteins, leading to novel bioactive materials. The wide substrate range of the BioF tag presumably enables protein immobilization in vitro of virtually all natural PHAs as well as blends, copolymers, or artificial chemically modified derivatives with novel physicochemical properties. Moreover, the strength of protein adsorption may be easily modulated by varying the coating of the support, providing new perspectives for the engineering of bioactive materials that require a tight control of protein loading.
|Reproducció del document publicat a: https://doi.org/10.1128/AEM.02595-17
|It is part of:
|Applied and Environmental Microbiology, 2018, vol. 84, num. 4, p. e02595
|Appears in Collections:
|Articles publicats en revistes (Institut d'lnvestigació Biomèdica de Bellvitge (IDIBELL))
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