Effect of pyk gene location on constructing dATP conversion E. coli strain


A dATP conversion E. coli strain could be constructed when both pyk and adk1 gene were expressed successfully. pyk gene encodes pyruvate kinase (PK) could be expressed, when inserted it before adk1 gene which encodes adenylate kinase (AK) in plasmid pET-pyk-adk1 after transform into E. coli and the recombinant could be used to convert dATP from dAMP. Another plasmid pET-adk1-pyk, which inserted pyk gene behind of adk1, the recombinant E. coli transformed with this plasmid could not convert dAMP into dATP, pyk gene cannot be translated in this recombinant. The different translation levels of pyk with gene location switching caused mainly by the different secondary structures formed by the 5’-untranslation regions and the gene sequence of its5’-terminal. The dATP product E. coli strain could be constructed when cloned pyk gene at an optimum location.

Share and Cite:

Xin, X. and Bao, J. (2013) Effect of pyk gene location on constructing dATP conversion E. coli strain. Advances in Bioscience and Biotechnology, 4, 75-80. doi: 10.4236/abb.2013.41011.

Conflicts of Interest

The authors declare no conflicts of interest.


[1] Bao, J. and Ryu, D.D.Y. (2005) Biosynthesis reaction mechanism and kinetics of deoxynucleoside triphosphates dATP and dGTP. Biotechnology Bioengeering, 89, 485-491. doi:10.1002/bit.20380
[2] Smolke, C.D. and Keasling, J.D. (2002) Effect of gene location, mrna secondary structures, and rnase sites on expression of two genes in an engineered operon. Biotechnology Bioengeering, 80, 762-776. doi:10.1002/bit.10434
[3] Alifano, P., Bruni, C.B. and Carlomagno, M.S. (1994) Control of mRNA processing and decay in prokaryotes. Genetica, 94, 157-172. doi:10.1007/BF01443430
[4] Dieter, V., Manfred, W., Cordula, N., Sabine, W. and Bernd, B. (2004) Analyzing and enhancing mRNA translational efficiency in an Escherichia coli in vitro expression system. Biochemical and Biophysical Research Communications, 318, 601-614. doi:10.1016/j.bbrc.2004.04.064
[5] Ravanshad, M., Sabahi, F. and Mahboudi, F. (2007) Quantification analysis of dot blot assays for human immunodeficiency virus type 1 and 2 antibodies. Iranian Journal of Basic Medical, 10, 132-138.
[6] Zuker, M. (2003) Web server for nucleic acid folding and hybridization prediction. Nucleic Acids Research, 31, 3406-3415. doi:10.1093/nar/gkg595
[7] Kimura, S. and Iyanagi, T. (2003) High-level expression of porcine liver cytochrome P-450 reductase catalytic domain in Escherichia coli by modulating the predicted local secondary structure of mRNA. Journal of Biochemistry, 134, 403-413.
[8] Geissmann, T., Marzi, S. and Romby, P. (2009) The role of mRNA structure in translational control in bacteria. RNA Biology, 6, 153-160. doi:10.4161/rna.6.2.8047
[9] Pfleger, B. F., Fawzi, N. J. and Keasling, J.D. (2005) Optimization of DsRed production in Escherichia coli: effect of ribosome binding site sequestration on translation efficiency. Biotechnology Bioengeering, 92, 553-558. doi:10.1002/bit.20630
[10] Nilsson, G., Belasco, J. G., Cohen, S. N. and Von Gabain, A. (1987) Effect of premature termination of translation on messenger RNA stability depends on the site of ribosome release. Proceedings of the National Academy of Science, 84, 4890-4894. doi:10.1073/pnas.84.14.4890

Copyright © 2023 by authors and Scientific Research Publishing Inc.

Creative Commons License

This work and the related PDF file are licensed under a Creative Commons Attribution 4.0 International License.