Modification of the loops in the ligand-binding site turns avidin into a steroid-binding protein.
BMC Biotechnol
; 11: 64, 2011 Jun 09.
Article
em En
| MEDLINE
| ID: mdl-21658230
ABSTRACT
BACKGROUND:
Engineered proteins, with non-immunoglobulin scaffolds, have become an important alternative to antibodies in many biotechnical and therapeutic applications. When compared to antibodies, tailored proteins may provide advantageous properties such as a smaller size or a more stable structure.RESULTS:
Avidin is a widely used protein in biomedicine and biotechnology. To tailor the binding properties of avidin, we have designed a sequence-randomized avidin library with mutagenesis focused at the loop area of the binding site. Selection from the generated library led to the isolation of a steroid-binding avidin mutant (sbAvd-1) showing micromolar affinity towards testosterone (Kd ~ 9 µM). Furthermore, a gene library based on the sbAvd-1 gene was created by randomizing the loop area between ß-strands 3 and 4. Phage display selection from this library led to the isolation of a steroid-binding protein with significantly decreased biotin binding affinity compared to sbAvd-1. Importantly, differential scanning calorimetry and analytical gel-filtration revealed that the high stability and the tetrameric structure were preserved in these engineered avidins.CONCLUSIONS:
The high stability and structural properties of avidin make it an attractive molecule for the engineering of novel receptors. This methodology may allow the use of avidin as a universal scaffold in the development of novel receptors for small molecules.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Testosterona
/
Avidina
Tipo de estudo:
Clinical_trials
Idioma:
En
Revista:
BMC Biotechnol
Assunto da revista:
BIOTECNOLOGIA
Ano de publicação:
2011
Tipo de documento:
Article
País de afiliação:
Finlândia