Molecular and in silico characterization of cold shock proteins (CSP2 and CSP5) in polar isolate Pseudomonas fragi AC

Despite the difficult ecological conditions of Antarctica, the largest component of biomass capable of surviving in the region is prokaryotic organisms and they have special adaptation mechanisms. Microorganisms that can adapt to these extreme conditions are potential gene sources for many biomolecules, from the mineralization of pollutants to cold inducible genes and extremozymes. The aim of this study is to determine the cold shock proteins in the polar isolate, psychrotolerant Pseudomonas fragi AC, and to analyze the expressions of the genes encoding these proteins at different temperature values by qPCR for the first time in the literature. A total of 10 genes were determined in the present database and the representative two genes were identified here. The AC-csp2 and AC-csp5 fragments were cloned by PCR and sequenced. According to the results of the sequence analysis, AC-csp2 was 100% identical with its counterparts in the literature, while AC-csp5 showed a difference of 1 amino acid in its protein structure, despite the 17-base difference in the nucleotide sequence. The identity of the csp2 and csp5 with each other was determined as 73%. in silico characterization of the proteins showed that both genes interact closely with ATP-dependent helicase. It was observed that both proteins had 54% β-strand formation and no α-helix formation. AC-CSP2 contains 100% the same domain as CapA, the cold shock protein of P. fluorescens, while AC-CSP5 contains 98.4% the same domain as the temperature acclimation protein, the product of tapB gene, in P. fragi. According to qPCR analysis, the expression of csp2 increased at 28 °C (p<0.05) compared to that at 15 °C, and the expression of csp5 gene showed a statistically significant decrease at 28 °C. Accordingly, the relative expression value of the csp5 gene compared to the control was 0.004 at 28 °C (p<0.0001). Within the scope of the study, the 16S rRNA sequence of P. fragi AC strain was registered in NCBI/GenBank with the accession number OR150484 and the csp2 and csp5 genes with the related accession numbers. In future studies, it is aimed to investigate the role of heterologous expression of P. fragi csp2 and csp5 genes in tolerance to cold stress in different strains.