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Journal of Bacteriology, January 2008, p. 718-726, Vol. 190, No. 2
0021-9193/08/$08.00+0     doi:10.1128/JB.01020-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Characterization of Nucleotide Pools as a Function of Physiological State in Escherichia coli{triangledown}

Michael H. Buckstein,1 Jian He,2 and Harvey Rubin1*

Division of Infectious Diseases,1 Department of Chemistry, University of Pennsylvania, Philadelphia, Pennsylvania 191042

Received 27 June 2007/ Accepted 17 October 2007

Using a modified method that involves minimal manipulation of cells, we report new information about nucleotide pool sizes and changes throughout the Escherichia coli growth curve. Nucleotide pool sizes are critically dependent on sample manipulation and extraction methods. Centrifugation and even short (2 min) lapses in sample preparation can dramatically affect results. The measured ATP concentration at three different growth rates is at least 3 mM, well above the 0.8 mM needed to saturate the rRNA promoter P1 in vitro. Many of the pools, including ATP, GTP, and UTP, begin to decrease while the cells are still in mid-log growth. After an almost universal drop in nucleotide concentration as the cells transition from logarithmic to stationary phase, there is a "rebound" of certain nucleotides, most notably ATP, after the cells enter stationary phase, followed by a progressive decrease. UTP, in contrast, increases as the cells transition into stationary phase. The higher UTP values might be related to elevated UDP-glucose/galactose, which was found to be at higher concentrations than expected in stationary phase. dTTP is the most abundant deoxynucleoside triphosphate (dNTP) in the cell despite the fact that its precursors, UDP and UTP, are not. All dNTPs decrease through the growth curve but do not have the abrupt drop, as seen with other nucleotides when the cells transition into stationary phase.

* Corresponding author. Mailing address: Division of Infectious Diseases, Department of Medicine, 522 Johnson Pavilion, 36th and Hamilton Walk, University of Pennsylvania, Philadelphia, PA 19104. Phone: (215) 662-6475. Fax: (215) 573-1240. E-mail: rubinh{at}mail.med.upenn.edu

{triangledown} Published ahead of print on 26 October 2007.

Journal of Bacteriology, January 2008, p. 718-726, Vol. 190, No. 2
0021-9193/08/$08.00+0     doi:10.1128/JB.01020-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.



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