Structural and Dynamic Features of Cold-Shock Proteins of Listeria monocytogenes, a Psychrophilic Bacterium

Abstract

Cold-shock proteins (Csps), proteins expressed when the ambient temperature drops below the growth-supporting temperature, bind to single-stranded nucleic acids and act as RNA chaperones to regulate translation. Listeria monocytogenes is a psychrophilic food-borne pathogen that is problematic for the food industry. Structures of Csps from psychrophilic bacteria have not yet been studied. Despite dramatic differences in the thermostability of Csps of various thermophilic microorganisms, these proteins share a high degree of primary sequence homology and a high degree of three-dimensional structural similarity. Here, we investigated the structural and dynamic features as well as the thermostability of L. monocytogenes CspA (Lm-CspA). Lm-CspA has a five-stranded β-barrel structure with hydrophobic core packing and two salt bridges. When heptathymidine (dT(7)) binds, values for the heteronuclear nuclear Overhauser effect and order parameters of residues in surface loop regions near nucleic acid binding sites increase dramatically. Moreover, Carr-Purcell-Meiboom-Gill experiments showed that slow motions observed for the nucleic acid binding residues K7, W8, F15, F27, and R56 disappeared in Lm-CspA-dT(7). Lm-CspA is less thermostable than mesophilic and thermophilic Csps, with a lower melting temperature (40 °C). The structural flexibility that accompanies longer surface loops and less hydrophobic core packing and a number of salt bridges and unfavorable electrostatic repulsion are likely key factors in the low thermostability of Lm-CspA. This implies that the large conformational flexibility of psychrophilic Lm-CspA, which more easily accommodates nucleic acids at low temperature, is required for RNA chaperone function under cold-shock conditions and for the cold adaptation of L. monocytogenes.

RNA binding and chaperone activity of theE. colicold-shock protein CspA

“…For E. coli CspA, we have demonstrated here that under temperature conditions (45°C) where part of the protein is unfolded, binding to an RNA hairpin shifts the population equilibrium toward the folded state. Similar observations have been made for B. subtilis CspB (18) as well as for L. monocytogenes CspA (14) upon interaction with single-stranded DNA. We also have some indication that RNA binding is accompanied by a loss in local entropy at the binding interface.…”

Novel Structural Components Contribute to the High Thermal Stability of Acyl Carrier Protein from Enterococcus faecalis

“…In previous studies, Ile 54 in Ec-ACP, which corresponds to Ile 57 in Ef-ACP, shows a large chemical shift perturbation in the ACP spectra by the addition of FabA (65). Here, conformational exchange in Ile 57 and Ser 58 in the ␣ 2 ␣ 3 loop was observed for the first time, and the flexibility of the ␣ 2 ␣ 3 loop might play an important role in its interaction with other FAS enzymes that could promote release of the growing acyl moieties from the hydrophobic pocket of ACP.…”

“…The recycle delay and proton saturation times in the hNOE measurement were 4.5 and 3.0 s, respectively. All relaxation spectra were acquired and analyzed as described in our previous reports (53,54). hNOE was calculated from the ratio of peak heights with and without proton saturation pulses.…”

Microbial Diversity in Extreme Marine Habitats and Their Biomolecules

“…Taxonomically, cold-loving and cold tolerant microbes are found in both Archaea and Bacteria domains and they are distributed over numerous genera, such as Arthrobacter , Colwellia , Exiguobacterium [111], Gelidibacter , Glaciecola [112], Halobacillus , Halomonas [113], Hyphomonas , Listeria [114], Marinobacter [115], Methanococcoides , Methanogenium [116], Moritella [117], Planococcus [118], Pseudoalteromonas , Pseudomonas [119], Psychrobacter , Psychroflexus [120], Psychromonas [104], Psychroserpens [100], Shewanella [121] and Sphingomonas (Table 4). …”