RT Journal Article
T1 Dissecting the Polyhydroxyalkanoate-binding domain of the PhaF Phasin: rational design of a minimized affinity tag
A1 Mato, Aranzazu
A1 Blanco, Francisco  G.
A1 Maestro García-Donas, María Beatriz
A1 Sanz, Jesús M.
A1 Pérez-Gil, Jesús
A1 Prieto, M. Auxiliadora
AB Phasin PhaF from Pseudomonas putida consists of a modular protein whose N-terminal domain (BioF) has been demonstrated to be responsible for binding to the polyhydroxyalkanoate (PHA) granule. BioF has been exploited for biotechnological purposes as an affinity tag in the functionalization of PHA beads with fusion proteins both in vivo and in vitro. The structural model of this domain suggests an amphipathic -helical conformation with the hydrophobic residues facing the PHA granule. In this work, we analyzed the mean hydrophobicity and the hydrophobic moment of the native BioF tag to rationally design shorter versions that maintain affinity for the granule. Hybrid proteins containing the green fluorescent protein (GFP) fused to the BioF derivatives were studied for in vivo localization on PHA, stability on the surface of the PHA granule against pH, temperature, and ionic strength, and their possible influence on PHA synthesis. Based on the results obtained, a minimized BioF tag for PHA functionalization has been proposed (MinP) that retains similar binding properties but possesses an attractive biotechnological potential derived from its reduced size. The MinP tag was further validated by analyzing the functionality and stability of the fusion proteins MinP–-galactosidase and MinP-CueO from Escherichia coli.
PB American Society for Microbiology
SN 0099-2240, ESSN: 1098-5336
YR 2020
FD 2020-06-02
LK https://hdl.handle.net/20.500.14352/6323
UL https://hdl.handle.net/20.500.14352/6323
LA eng
NO Unión Europea. Horizonte 2020
NO Ministerio de Ciencia e Innovación (MICINN)
NO Comunidad de Madrid
DS Docta Complutense
RD 8 abr 2025