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

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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.

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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

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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.

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Authors and Affiliations

  1. ICAR-National Bureau of Agriculturally Important Microorganisms (NBAIM), Mau, Uttar Pradesh, 275103, India

    Srikant Awasthi, Anjney Sharma, Pragya Saxena, Jagriti Yadav, K. Pandiyan, M. Kumar, Arjun Singh, Hillol Chakdar, Alok K. Srivastava & Anil K. Saxena

  2. ICAR-Indian Institute of Wheat and Barley Research (IIWBR), Karnal, India

    Prem L. Kashyap

  3. ICAR-Indian Agricultural Statistics Research Institute (IASRI), New Delhi, 110012, India

    Arpan Bhowmik

Authors
  1. Srikant Awasthi
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  2. Anjney Sharma
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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.

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Correspondence to Hillol Chakdar.

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Supplementary Figure 1

Multiple sequence alignment of cspA sequences from different Pseudomonas strains (JPEG 374 kb)

Supplementary Figure 2

The secondary structure of CSP from Pseudomonas koreensis P2 (JPEG 75 kb)

Supplementary Figure 3

Multiple alignment of the deduced amino acid sequences of CspA of Pseudomonas koreensisP2 (JPEG 467 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)

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)

Supplementary Figure 6

Verify 3D score of predicted CspA model (JPEG 77 kb)

Supplementary Figure 7

Overall quality factor for CspA protein model obtained from ERRAT server (JPEG 14 kb)

Supplementary Figure 8

Binding pockets (shown in different colors) of CspA protein from Psedomonas koreensis P2 (JPEG 58 kb)

Supplementary Figure 9

Figure showing electrostatic potential on CspA model (JPEG 54 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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Awasthi, S., Sharma, A., Saxena, P. et al. Molecular detection and in silico characterization of cold shock protein coding gene (cspA) from cold adaptive Pseudomonas koreensis. J. Plant Biochem. Biotechnol. 28, 405–413 (2019). https://doi.org/10.1007/s13562-019-00500-8

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