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Molecular detection and in silico characterization of cold shock protein coding gene (cspA) from cold adaptive Pseudomonas koreensis

  • Srikant Awasthi
  • Anjney Sharma
  • Pragya Saxena
  • Jagriti Yadav
  • K. Pandiyan
  • M. Kumar
  • Arjun Singh
  • Hillol ChakdarEmail author
  • Arpan Bhowmik
  • Prem L. Kashyap
  • Alok K. Srivastava
  • Anil K. Saxena
Original Article
  • 25 Downloads

Abstract

Cold shock proteins (CSPs) are greatly conserved family of structurally related DNA binding proteins which are produced during temperature drift. A 213 bp long cspA gene was cloned and sequenced from Pseudomonas koreensis P2 in the present study. The expression analysis of the cspA showed > 2.5 folds increase in the mRNA level at 15 °C while the expression was almost on par at 30 °C and 5 °C indicating its role in moderately low temperature. In silico analyses of the gene showed that the gene codes for 7.69 kDa protein which was phylogenetically very similar to CspA present in Pseudomonads. Amino acid composition of the CspA from P. koreensis was different from that of mesophilic Pseudomonas and tiny/small amino varied significantly between CspA of cold adaptive and mesophilic species. The CspA from P. koreensis P2 contained RNP motifs involved in binding of DNA and RNA. Phylogenetic analyses revealed that the CspA protein of P. koreensis P2 was more close to CspA of distant subgroups of Pseudomonas like P. fluorescens and P. putida subgroup indicating a possible intra-specific gene transfer.

Keywords

Cold shock protein Pseudomonas koreensis Psychrophiles Gene expression Molecular modeling 

Abbreviations

CSPs

Cold shock proteins

GRAVY

Grand average hydropathy

MEGA

Molecular evolutionary genetics analysis

PDB

Protein Data Bank

NCBI

National Center for Biotechnology Information

NABPs

Nucleic acid binding proteins

DDBJ

DNA Data Bank of Japan

PMDB

Protein model database

C-score

Correlation scoring

MIQE

Minimum information for publication of quantitative RT PCR experiments

Notes

Acknowledgements

The authors gratefully acknowledge the financial assistance under network project ‘Application of Microorganisms in Agriculture and Allied Sectors (AMAAS)’ and “CRP Genomics” from Indian Council of Agricultural Research (ICAR), India.

Author contributions

HC conceptualized the study. Primers were designed by KM. Molecular works were carried out by ASh and PS. SA performed all computational analyses. JY performed the gene expression experiment. AB performed statistical analyses. HC, KM, KP and ASi drafted and revised the manuscript. AKS, PLK and AKS helped in execution of the experiments.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

13562_2019_500_MOESM1_ESM.jpg (375 kb)
Supplementary Figure 1 Multiple sequence alignment of cspA sequences from different Pseudomonas strains (JPEG 374 kb)
13562_2019_500_MOESM2_ESM.jpg (76 kb)
Supplementary Figure 2 The secondary structure of CSP from Pseudomonas koreensis P2 (JPEG 75 kb)
13562_2019_500_MOESM3_ESM.jpg (468 kb)
Supplementary Figure 3 Multiple alignment of the deduced amino acid sequences of CspA of Pseudomonas koreensisP2 (JPEG 467 kb)
13562_2019_500_MOESM4_ESM.jpg (85 kb)
Supplementary Figure 4 Ramchandran plot of the CspA model. The most favored regions are colored red, additional allowed, generously allowed and disallowed regions are indicated as yellow, light yellow and white fields, respectively (JPEG 84 kb)
13562_2019_500_MOESM5_ESM.jpg (71 kb)
Supplementary Figure 5 Model quality estimation plot obtained by QMEAN server. The area built by the circles colored in different shades of gray in the plot represents the QMEAN scores of the reference structures from the PDB (JPEG 70 kb)
13562_2019_500_MOESM6_ESM.jpg (78 kb)
Supplementary Figure 6 Verify 3D score of predicted CspA model (JPEG 77 kb)
13562_2019_500_MOESM7_ESM.jpg (14 kb)
Supplementary Figure 7 Overall quality factor for CspA protein model obtained from ERRAT server (JPEG 14 kb)
13562_2019_500_MOESM8_ESM.jpg (58 kb)
Supplementary Figure 8 Binding pockets (shown in different colors) of CspA protein from Psedomonas koreensis P2 (JPEG 58 kb)
13562_2019_500_MOESM9_ESM.jpg (54 kb)
Supplementary Figure 9 Figure showing electrostatic potential on CspA model (JPEG 54 kb)
13562_2019_500_MOESM10_ESM.jpg (49 kb)
Supplementary Figure 10 Three largest positive patches (in different blue color balls), calculated on a structural model of CspA. The model is predicted to be NA-binding (JPEG 49 kb)

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Copyright information

© Society for Plant Biochemistry and Biotechnology 2019

Authors and Affiliations

  • Srikant Awasthi
    • 1
  • Anjney Sharma
    • 1
  • Pragya Saxena
    • 1
  • Jagriti Yadav
    • 1
  • K. Pandiyan
    • 1
  • M. Kumar
    • 1
  • Arjun Singh
    • 1
  • Hillol Chakdar
    • 1
    Email author
  • Arpan Bhowmik
    • 3
  • Prem L. Kashyap
    • 2
  • Alok K. Srivastava
    • 1
  • Anil K. Saxena
    • 1
  1. 1.ICAR-National Bureau of Agriculturally Important Microorganisms (NBAIM)MauIndia
  2. 2.ICAR-Indian Institute of Wheat and Barley Research (IIWBR)KarnalIndia
  3. 3.ICAR-Indian Agricultural Statistics Research Institute (IASRI)New DelhiIndia

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