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Extremophile Protein Database
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Psychrophile

Psychrophiles are cold-adapted microorganisms which have an optimal temperature for growth at about 20° C or lower. Cold adaptations of these microorganisms challenges include: decreased membrane fluidity; altered transport of nutrients and waste products; decreased rates of transcription, translation and cell division; reduced enzyme activity; protein cold-denaturation; inappropriate protein folding; and intracellular ice formation. However, recent developments based on cold-adapted organisms and on their biomolecules, have clearly demonstrated the huge potential of psychrophiles. They have useful applications in various domains such as molecular biology, medical research, industrial food or feed technologies, detergents or cosmetics (Margesinand Feller, 2010).

Factors responsible for Psychrostability

Genomic adaptations

A large repertoire of psychrophilic organisms has been identified. Search for genomes with the key words "psychrophilic" in NCBI genome search results in a total of 14 hits for psychrophiles. The five complete and more focused genomes are those from Desulfotalea psychrophila (Rabus et al, 2004), Colwellia psychrerythraea 34H (Methe et al, 2005), Pseudoalteromonas haloplanktis TAC125 (Medigue et al, 2005), Methanogenium frigidum and Methanococcoides burtonii (Saunders et al, 2003). As expected, several cold shock proteins and proteins that are involved in unsaturated fatty-acid synthesis have been identified in these genomes. In addition to the classical lipid desaturases, two gene clusters are possibly involved in membrane rigidity/fluidity through the degradation of steroids or hopanoids, have been found in the P. haloplanktis genome (D'Amico et al. 2006). At low temperatures, the solubility of gases and the production of toxic reactive oxygen species (ROS) increase significantly. To counteract this, C. psychrerythraea and D. psychrophila have an enhanced antioxidant capacity owing to the presence of several genes that encode catalases and superoxide dismutases (Podar and Reysenbach, 2006). Psychrophilic bacteria also produce nucleic acid-binding proteins such as RNA helicase to relieve strong interactions between DNA strands and secondary structures in RNA which impair transcription, translation, and replication (Feller and Charles, 2003).

Proteomic adaptations

The psychrophilic stability can be called as the main branch of extreme adaptations as thermophilic and halophilic stability whereas alkaliphilic, acidophilic and barophilic intersect these branches (Reed et al. 2013). It is also interesting to note that thermophiles and psychrophiles choose features opposite to each other for enhancing their stability. For example, thermostability increases by increase in protein rigidity whereas psychrophilicity increases by increase in flexibility. Another complication in understanding extreme-stability is that certain proteins for example; lipases from psychrophilic fungus Candida antarctica which is supposed to be psychrophilic is actually thermostable. They are stable due to reduced rigidity and higher number of Van der Waals forces, reduction in electrostatic interaction and increase in hydrogen bonding (D'Amico et al. 2006). An increase in the flexibility of proteins is the key to low-temperature adaptation. The protein stability at lower temperatures favour protein flexibility than the protein rigidity at higher temperature (Pack and Yoo 2004). This is achieved by decreasing the number of Pro and Arg residues (which leads to the rigidity of proteins by restricting backbone rotations) and increasing the number of Gly residues in their sequences (Feller and Gerday 2003).While comparing the psychrophilic proteins with mesophilic proteins, they also comprise a significantly higher proportion of amino acids that contribute to higher protein flexibility in the coil regions of proteins, such as those with tiny/small or neutral side chains (Creighton 1997; Metpally and Reddy 2009). Metapally and Reddy (2009) also said that in psychrophilic proteins, Ser, Asp, Thr and Ala are overrepresented in the coil regions of secondary structures, whilst Glu and Leu are underrepresented in the helical regions. Comparing the amino acid properties, the psychrophilic proteins with aliphatic, basic, aromatic and hydrophilic side chains are underrepresented in the helical regions of proteins of psychrophiles (Metpally and Reddy 2009).

The structural features of psychrophilic proteins have not been reported due to lack of 3D-structure. Alternatively, homology modeling may be employed to determine the structural features of proteins from psychrophilic microorganisms by comparative analysis of mesophilic and thermophilic proteins. However, the interpretation of hydrogen bonds in homology models is not much reliable because hydrogen bonds are highly directional. No hydrogen bond differences were identified from comparative analyses of the X-ray structures of α-amylase, citrate synthase, and malate dehydrogenase (Russell et al. 1998). The total difference in stabilization energy may be as little as 40-50 kJ mol/1 between homologs from a psychrophile and mesophile, it is clear that this may be accounted for by a few critically placed hydrogen bonds. A decrease in the trend of number of hydrogen bonds at domain interfaces in psychrophiles as compared to mesophilic counterpart was reported by Feller et al. (1997), Marshall, (1997) and Russell et al. (1998). For example, cold adaptation of a triosephosphate isomerase was linked to an Ala replacement of a Ser; Ser is expected to confer thermostability by forming two additional intramolecular hydrogen bonds (Alvarez et al. 1998).

Several comparative studies based on X-ray structure data have reported about the flexibility of psychrophilic proteins in their native state, thus they require lower number of electrostatic interactions and salt bridges under cold-adapted conditions. For example, cold adapted subtilisin has two salt bridges compared with five and ten salt bridges in the homologs from mesophiles and thermophiles, respectively (Davail et al. 1994). Similarly, Kim et al. in 1999 said that, a reduced number of inter-subunit and ion-pair networks also appear to be important for the heat lability of a malate dehydrogenase from a psychrophile (Kim et al. 1999).

Psychrophilic proteins enhance their flexibility by decreasing the number and strength of various interactions (Gianese et al. 2001; Violot et al. 2005). In particular, there is a significant decrease in the interaction between hydrophobic residues and between hydrophobic and aromatic residues, as would be expected given the temperature dependence of the hydrophobic effect (Goldstein 2007). Unlike thermophilic proteins, the psychrophilic proteins are flexible in nature, thus lack aromatic interactions in them (Davail et al. 1994). For example, heat-labile subtilisin, from the Antarctic psychrophile Bacillus TA41 has a general lack of aromatic interactions in contrast to 11 interactions identified on the surface of a thermophilic homolog, subtilisin Carlsberg from Bacillus licheniformis (Davail et al. 1994). Similarly, in a β-lactamase of Antarctic psychrophile Psychrobacter immobilis A5 lack aromatic interactions when compared with homologous mesophilic counterpart (Feller et al. 1997).

The molecular chaperones, cold-shock proteins assist in protein renaturation during the growth of psychrophiles (Berger et al. 1996). The presence of unique antifreeze proteins such as the proteins from Marinomonas primoryensis and Pseudomonas putida GR12-2 lower the freezing point of cellular water (Feller and Gerday 2003). On the proteome level, thermophiles and psychrophiles differ markedly from mesophiles by synthesizing chaperones and antifreeze proteins, respectively, which leads to refolding of denatured proteins at such extremes.

Further Reading

Alvarez M et al. (1998) Triose-phosphate Isomerase (TIM) of the Psychrophilic Bacterium Vibrio marinus kinetic and structural properties Journal of Biological Chemistry 273:2199-2206.

Berger F, Morellet N, Menu F, Potier P (1996) Cold shock and cold acclimation proteins in the psychrotrophic bacterium Arthrobacterglobiformis SI55 Journal of bacteriology 178:2999-3007.

Creighton TE (1997) Protein structure: a practical approach

D'Amico S, Collins T, Marx JC, Feller G, Gerday C (2006) Psychrophilic microorganisms: challenges for life EMBO reports 7:385-389.

Davail S, Feller G, Narinx E, Gerday C (1994) Cold adaptation of proteins. Purification, characterization, and sequence of the heat-labile subtilisin from the antarcticpsychrophile Bacillus TA41 Journal of Biological Chemistry 269:17448-17453.

Feller G, Arpigny J, Narinx E, Gerday C (1997) Molecular adaptations of enzymes from psychrophilic organisms Comparative Biochemistry and Physiology Part A: Physiology 118:495-499.

Feller G, Gerday C (2003) Psychrophilic enzymes: hot topics in cold adaptation Nature reviews microbiology 1:200-208.

Gianese G, Argos P, Pascarella S (2001) Structural adaptation of enzymes to low temperatures Protein engineering 14:141-148.

Kim S-Y, Hwang KY, Kim S-H, Sung H-C, Han YS, Cho Y (1999) Structural basis for cold adaptation sequence, biochemical properties, and crystal structure of malate dehydrogenase from a psychrophileAquaspirilliumarcticum Journal of Biological Chemistry 274:11761-11767.

Margesin R, Feller G (2010) Biotechnological applications of psychrophilesEnvironmental technology31(8-9), 835-844.

Marshall CJ (1997) Cold-adapted enzymes Trends in biotechnology 15:359-364.

Medigue C et al. (2005) Coping with cold: the genome of the versatile marine Antarctica bacterium Pseudoalteromonashaloplanktis TAC125. Genome Res15: 1325-1335.

Methe BA et al. (2005) The psychrophilic lifestyle as revealed by the genome sequence of Colwelliapsychrerythraea 34H through genomic and proteomic analyses. ProcNatlAcadSci USA102: 10913-10918.

Metpally RPR, Reddy BVB (2009) Comparative proteome analysis of psychrophilic versus mesophilic bacterial species: Insights into the molecular basis of cold adaptation of proteins BMC genomics 10:1.

Pack SP, Yoo YJ (2004) Protein thermostability: structure-based difference of amino acid between thermophilic and mesophilic proteins Journal of Biotechnology 111:269-277.

Podar M, Reysenbach AL (2006) New opportunities revealed by biotechnological explorations of extremophiles Current opinion in biotechnology, 17(3), 250-255.

Rabus R et al. (2004) The genome of Desulfotaleapsychrophila, a sulfate-reducing bacterium from permanently cold Arctic sediments. Environ Microbiol 6: 887-902.

Reed CJ, Lewis H, Trejo E, Winston V, Evilia C (2013) Protein adaptations in archaeal extremophiles Archaea 2013.

Russell NJ (1998) Molecular adaptations in psychrophilic bacteria: potential for biotechnological applications In Biotechnology of Extremophiles(pp. 1-21). Springer Berlin Heidelberg.

Russell RJ, Gerike U, Danson MJ, Hough DW, Taylor GL (1998) Structural adaptations of the cold-active citrate synthase from an Antarctic bacterium Structure 6:351-361.

Saunders NF et al. (2003) Mechanisms of thermal adaptation revealed from the genomes of the Antarctic ArchaeaMethanogeniumfrigidum and Methanococcoidesburtonii. Genome Res13: 1580-1588.

Violot S et al. (2005) Structure of a full length psychrophilic cellulase from Pseudoalteromonashaloplanktis revealed by X-ray diffraction and small angle X-ray scattering Journal of molecular biology 348:1211-1224.