Proteomic analysis of Psychrobacter cryohalolentis K5 during growth at subzero temperatures

Abstract

It is crucial to examine the physiological processes of psychrophiles at temperatures below 4 degrees C, particularly to facilitate extrapolation of laboratory results to in situ activity. Using two dimensional electrophoresis, we examined patterns of protein abundance during growth at 16, 4, and -4 degrees C of the eurypsychrophile Psychrobacter cryohalolentis K5 and report the first identification of cold inducible proteins (CIPs) present during growth at subzero temperatures. Growth temperature substantially reprogrammed the proteome; the relative abundance of 303 of the 618 protein spots detected (approximately 31% of the proteins at each growth temperature) varied significantly with temperature. Five CIPs were detected specifically at -4 degrees C; their identities (AtpF, EF-Ts, TolC, Pcryo_1988, and FecA) suggested specific stress on energy production, protein synthesis, and transport during growth at subzero temperatures. The need for continual relief of low-temperature stress on these cellular processes was confirmed via identification of 22 additional CIPs whose abundance increased during growth at -4 degrees C (relative to higher temperatures). Our data suggested that iron may be limiting during growth at subzero temperatures and that a cold-adapted allele was employed at -4 degrees C for transport of iron. In summary, these data suggest that low-temperature stresses continue to intensify as growth temperatures decrease to -4 degrees C.

Life in the Cold: Proteomics of the Antarctic Bacterium Pseudoalteromonas haloplanktis

“…The genome of P. haloplanktis also reveals several insights into ROS protection such as deletion of ROS producing pathways, several occurrences of dioxygenases and the repair mechanisms of oxidized compounds (Medigue et al, 2005). Similar observations have been made in the genome and proteome of other psychrophilic microorganisms (Rabus et al, 2004;Medigue et al, 2005;Methe et al, 2005;Duchaud et al, 2007;Bakermans et al, 2007;Ayub et al, 2009;Piette et al, 2010), revealing a general constraint on these bacteria. However, we found that at 18 °C, another type of oxidative stress is induced by stimulation of metabolic activity.…”

“…Cold shock proteins that have been identified as CAPs in P. haloplanktis include Pnp (+4x), TypA (+5x) and the trigger factor TF (+38x) that are involved in distinct functions (degradosome, membrane integrity and protein folding, respectively). Sustained synthesis of various cold shock protein-homologues has been also reported in other cold-adapted bacteria (Bakermans et al, 2007;Kawamoto et al, 2007;Bergholz et al, 2009). There are therefore striking similarities between the cold shock response in mesophiles and cold adaptation in psychrophiles.…”

“…On the other hand, the second group of proteins that displays the highest repression factors at 4 °C is represented by the oxidative stress-related proteins catalase (-6.5x), glutathione reductase (-8.1x) and peroxiredoxin (-15.7x). At first sight, this may be regarded as a conflicting result because conclusive evidences have indicated that psychrophiles are exposed to a permanent oxidative stress at low temperatures, which originates from improved dioxygen solubility and increased ROS stability (Rabus et al, 2004;Medigue et al, 2005;Methe et al, 2005;Duchaud et al, 2007;Bakermans et al, 2007;Ayub et al, 2009;Piette et al, 2010). In order to reconcile these apparent contradictions, it should be recalled that the general aerobic metabolism of the Antarctic bacterium is stimulated at 18 °C, also resulting in ROS production.…”

Psychrobacter arcticus 273-4 Uses Resource Efficiency and Molecular Motion Adaptations for Subzero Temperature Growth

“…Gene expression studies of both psychrotolerant and strict mesophiles growing at suboptimal temperatures have detected increased expression of proteins for destabilization of nucleic acid secondary structures, maintenance of membrane fluidity, chaperones, uptake of compatible solutes, and energy metabolism (2,36,47). However, reports of the global modifications to metabolism necessary for growth of psychrophiles at temperatures below 0°C remain rare (4,5). Analysis of the amino acid sequences and structures of psychrophilic enzymes has given rise to the flexibility concept, i.e., that a psychrophilic enzyme can exhibit increased catalytic activity at low temperature with limited loss of thermostability by adaptation for reduced numbers of stabilizing interactions between key amino acid residues (20,40).…”

“…For analysis of the proteome, cells were harvested from the remaining 475 ml of the transcriptome sample culture and analyzed as described by Bakermans et al (5). Briefly, samples were centrifuged at 10,000 ϫ g in a Sorvall RC-5B refrigerated centrifuge at the culture temperature for 10 min.…”