Chemical Evolution of a Bacterial Proteome - PubMed (original) (raw)

Chemical Evolution of a Bacterial Proteome

Michael Georg Hoesl et al. Angew Chem Int Ed Engl. 2015.

Abstract

We have changed the amino acid set of the genetic code of Escherichia coli by evolving cultures capable of growing on the synthetic noncanonical amino acid L-β-(thieno[3,2-b]pyrrolyl)alanine ([3,2]Tpa) as a sole surrogate for the canonical amino acid L-tryptophan (Trp). A long-term cultivation experiment in defined synthetic media resulted in the evolution of cells capable of surviving Trp→[3,2]Tpa substitutions in their proteomes in response to the 20,899 TGG codons of the E. coli W3110 genome. These evolved bacteria with new-to-nature amino acid composition showed robust growth in the complete absence of Trp. Our experimental results illustrate an approach for the evolution of synthetic cells with alternative biochemical building blocks.

Keywords: Escherichia coli; continuous evolution; genetic code translation; synthetic biology; tryptophan analogues.

© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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Figures

Figure 1

Figure 1

Structures of 1: tryptophan (Trp), 2: indole (Ind), 3: β-thieno[3,2_-b_]-pyrrole ([3,2]Tp), and L-β-(thieno[3,2_-b_]pyrrolyl)alanine ([3,2]Tpa).

Figure 2

Figure 2. Adaptation of E. coli strain CGSC# 7679 to [3,2]Tpa

The upper scale indicates the passage count while the lower scale shows the time in days. In each passage, OD600nm was measured. The growth temperature was decreased from 37°C to 30°C while going from NMM2s to NMM1. The different medium compositions are indicated on the right side and are described in details in section 4 in the SI.

Figure 3

Figure 3

Morphology and average distribution of cell length (A) and width (B) as measured by microscopy of MT0 and MT1. Strains were grown in NMM19 with either 100 μM of Indole (black dots → MT0; blue diamonds → MT1) or of [3,2]Tp (orange squares → MT1). Error bars represent the standard errors of the means of four experiments (with 350 cells studied by experiment). Inserts: examples of brightfield microscopy pictures of MT0 (0 μM [3,2]Tp /100 μM Indole) and MT1 (0 μM Indole/100 μM [3,2]Tp) cells grown in NMM19 and prepared on a 1% agarose-coated slide to be visualised by microscopy. The calibration bar indicates 5 μm. Details can be found in sections 9 and 10 in the SI.

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