Determination of amino acids misincorporation in recombinant protein by mass spectrometry

Authors

  • MZ Alam Department of Genetic Engineering and Biotechnology, Shahjalal University of Science and Technology, Sylhet
  • L Regioneiri RNA Biology Laboratory, Department of Biology and CESAM, University of Aveiro, Aveiro
  • MAS Santos RNA Biology Laboratory, Department of Biology and CESAM, University of Aveiro, Aveiro

DOI:

https://doi.org/10.3329/bjas.v42i1.15760

Keywords:

Amino acids misincorporation, mass spectrometry, recombinant protein, ?- galactosidase

Abstract

The synthesis of protein according to genetic code of a gene determines the basis of life and a stable proteome is necessary for cell homeostatis. However, errors occur naturally during translation of protein from its mRNA, which varies from 10-3 to 10-4 per codon. These errors are more frequent in recombinant protein overexpressed in heterologous hosts and affect protein functionality. The increasing amount of nonfunctional protein is often related to mistranslation of a gene under stress. In the present study, Saccharomyces cerevisiae as a host organism to overexpress E. coli lacZ gene fusion with GST to quantify misincorporation of amino acid in GST-? galactosidase recombinant protein. The yeast was treated with various stressors such as ethanol, chromium (CrO3), and aminoglycoside antibiotic - geneticin (G418) to induce protein aggregation. The misincorporation of amino acids was studied in soluble protein fractions by mass-spectrometry to determine how much misincorporation occur. We found that under experimental stress conditions the misincorporation of amino acids ranges from 5.6 ×10-3 to 8 × 10-3, which represents 60-80 fold higher than reported level.

DOI: http://dx.doi.org/10.3329/bjas.v42i1.15760

Bang. J. Anim. Sci. 2013. 42 (1): 11-19

Downloads

Download data is not yet available.
Abstract
151
PDF
298

Downloads

Published

2013-07-22

How to Cite

Alam, M., Regioneiri, L., & Santos, M. (2013). Determination of amino acids misincorporation in recombinant protein by mass spectrometry. Bangladesh Journal of Animal Science, 42(1), 11–19. https://doi.org/10.3329/bjas.v42i1.15760

Issue

Vol. 42 No. 1 (2013)

Section

Articles

License

© Bangladesh Journal of Animal Science.

Authors are required to transfer their copyright to the Bangladesh Journal of Animal Sciences.

All rights reserved. No parts of this publication may be reproduced, stored in a retrieval system or transmitted in any form or by any means, electronic, mechanical, photocopying, recording or otherwise without the prior permission of the Bangladesh Journal of Animal Science.