Abstract
The structural analyses of four metabolic enzymes that maintain and regulate the stationary growth phase of Escherichia coli have been performed primarily drawing on the results obtained from solution small angle X-ray scattering (SAXS) and other structural techniques. The proteins are (i) class I fructose-1,6-bisphosphate aldolase (FbaB); (ii) inorganic pyrophosphatase (PPase); (iii) 5-keto-4-deoxyuronate isomerase (KduI); and (iv) glutamate decarboxylase (GadA). The enzyme FbaB, that until now had an unknown structure, is predicted to fold into a TIM-barrel motif that form globular protomers which SAXS experiments show associate into decameric assemblies. In agreement with previously reported crystal structures, PPase forms hexamers in solution that are similar to the previously reported X-ray crystal structure. Both KduI and GadA that are responsible for carbohydrate (pectin) metabolism and acid stress responses, respectively, form polydisperse mixtures consisting of different oligomeric states. Overall the SAXS experiments yield additional insights into shape and organization of these metabolic enzymes and further demonstrate the utility of hybrid methods, i.e., solution SAXS combined with X-ray crystallography, bioinformatics and predictive 3D-structural modeling, as tools to enrich structural studies. The results highlight the structural complexity that the protein components of metabolic networks may adopt which cannot be fully captured using individual structural biology techniques.
Highlights
The transition of bacterial cells to the stationary phase of growth may result in lowered metabolic activity combined with an increase in resistance to stress effects caused by nutrient deprivation [1]
Concentration dependent effects on the measured scattering intensities were not observed for FbaB, keto 4-deoxyuronate isomerase (KduI) and glutamate decarboxylase (GadA) samples in the standard Tris buffer spanning 1.4–10.9 mg/ml (S1 Fig), whereas slight repulsive interparticle interference are noted at low-s in the small angle X-ray scattering (SAXS) data for PPase when the sample concentration is increased from 2.8–10.9 mg/ml, for s < 0.54 nm-1
When SAXS data are recorded from a sample without NaCl present, the Rg decreases from 4.8 nm to 4.4 nm, while the I(0) and average molecular masses (MM) estimate increases, suggesting significant structural rearrangements occur in the protein in response to a change in sample environment (Table 1; Fig 2B and 2C and see below)
Summary
The transition of bacterial cells to the stationary phase of growth may result in lowered metabolic activity combined with an increase in resistance to stress effects caused by nutrient deprivation [1]. Many cellular processes undergo changes during stationary phase or are induced/ activated in order to provide protective mechanisms against unfavorable environmental. Solution Structures of Four E. coli Proteins Involved in the Metabolism of Stationary-Phase Cells
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