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Changes in the Functional Activity of Phi11 Cro Protein is Mediated by Various Ions

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Abstract

Phi11, a temperate bacteriophage of Staphylococcus aureus, has been found to harbor a cro repressor gene which facilitates Phi11 to adopt the lytic mode of development. The Cro protein has been found to bind very specifically to a 15-bp operator DNA, located in the Phi11 cI–cro intergenic region [1]. To investigate the effects exerted by different ions upon the interaction between Cro and its cognate operator DNA, we have employed gel shift assays as well as circular dichroism spectral analysis. In this communication, we have shown that NH4 + and acetate− ions better facilitated the binding of Cro with its cognate operator as compared to Na+, K+ and Li+. Interestingly, Mg2+, carbonate2− and Citrate3− have an inhibitory effect upon the binding. The effect of the said ions upon the structure of Cro was also investigated by circular dichroism and it was found that other than Citrate3− ions, none of the other ions destabilised the protein. On the other hand, Mg2+ and carbonate2− ions maintained the structure of the protein but severely hampered its functional activity. Citrate3− ions severely unfolded Cro and also inhibited its function. Considering all the data, NH4 + and acetate− ions appeared to be more suitable in maintaining the biological activity of Cro.

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Abbreviations

bp:

Base pairs

O DNA:

cI–cro intergenic region bearing the cognate operator DNA for Phi11 Cro

His-Cro:

Phi11 Cro carrying 6-histidine-tag at its N-terminal end

SDS-PAGE:

Sodium dodecyl sulfate polyacrylamide gel electrophoresis

Ni–NTA:

Nickel–nitrilotriacetic acid

CD:

Circular dichroism

PCR:

Polymerase chain reaction

DMSO:

Dimethyl sulfoxide

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Acknowledgements

This work was supported by the financial assistance from BRNS [DAE, Govt. of India] [Sanction No. 2013/37B/19/BRNS] and DST-SERB [EMR/2015/002229] to Dr. Malabika Biswas. Mr. Avijit Das is a recipient of Junior Research Fellowship from BRNS [DAE, Govt. of India]. We are also thankful to CIF, UDSC New Delhi, India for carrying out CD spectra.

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  1. Department of Biological Sciences, BITS Pilani, K. K. Birla Goa Campus, NH17B, Zuarinagar, Goa, 403726, India

    Avijit Das & Malabika Biswas

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  1. Avijit Das
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  2. Malabika Biswas
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Correspondence to Malabika Biswas.

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Das, A., Biswas, M. Changes in the Functional Activity of Phi11 Cro Protein is Mediated by Various Ions. Protein J 35, 407–415 (2016). https://doi.org/10.1007/s10930-016-9684-8

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