- Poster Presentation
- Open Access
Performance of beta-galactosidase inclusion bodies in enzymatic bioprocesses
https://doi.org/10.1186/1475-2859-5-S1-P14
© García-Fruitós et al; licensee BioMed Central Ltd. 2006
- Published: 10 October 2006
Keywords
- Inclusion Body
- Soluble Fraction
- Capsid Protein
- Body Formation
- Recombinant Enzyme
Background
Inclusion body formation is a common event during bacterial over-expression of recombinant genes. This phenomenon represents a great matter of concern in biotechnology, because it has restricted the spectrum of proteins marketed in this field. In a previous work, we have observed that recombinant enzymes produced in bacteria are not completely inactivated when deposited as inclusion bodies [1] and that aggregation as inclusion bodies does not necessarily split protein population into active and inactive fractions. Therefore, we decided to further explore and fully characterize the behaviour of purified beta-galactosidase inclusion bodies in presence of substrate, during a small-scale bioprocess.
Results
A. Product formed by inclusion bodies (quadruplicate) through ONPG hydrolysis as determined at 414 nm. B. Control (inclusion bodies without ONPG).
:
Protein (%) | Inclusion bodies | Soluble | ||
---|---|---|---|---|
t2 min | t30 min | t2 min | t30 min | |
samples | 100 | 28 | <1 | 72 |
A. Product formed by soluble fraction (A) and inclusion bodies (B) through CPRG hydrolysis as determined at 540 nm.
Conclusion
We could conclude that, interestingly, when an enzyme aggregated as inclusion bodies is incubated with its substrate, part of this protein might be spontaneously solubilised in a process that seems to be eventually favoured by the presence of substrate. Moreover, this soluble protein shows considerable enzymatic activity that is a major contributor to the enzymatic process initiated by inclusion bodies.
Declarations
Acknowledgements
This work has been funded by BIO2004-00700 from MEC, Spain and 2005SGR-00956 (AGAUR). Elena García-Fruitós is recipient of a doctoral fellowship from MEC, Spain.
Authors’ Affiliations
References
- García-Fruitós E, González-Montalbán N, Morell M, Vera A, Ferraz RM, Arís A, Ventura S, Villaverde A: Aggregation as bacterial inclusion bodies does not imply inactivation of enzymes and fluorescent proteins. Microb Cell Fact. 2005, 4: 27- 10.1186/1475-2859-4-27.View ArticleGoogle Scholar
- Ferraz RM, Aris A, Villaverde A: Profiling the allosteric response of an engineered beta-galactosidase to its effector, anti-HIV antibody. Biochem Biophys Res Commun. 2004, 314: 854-860. 10.1016/j.bbrc.2003.12.169.View ArticleGoogle Scholar
Copyright
This article is published under license to BioMed Central Ltd.