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A single subunit MCM6 from pea forms homohexamer and functions as DNA helicase

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Abstract

The initiation of DNA replication starts from origins and is controlled by a multiprotein complex, which involves about twenty protein factors. One of the important factors is hetrohexameric minichromosome maintenance (MCM2-7) protein complex which is evolutionary conserved and functions as essential replicative helicase for DNA replication. Here we report the isolation and characterization of a single subunit of pea MCM protein complex, the MCM6. The deduced amino acid (827) sequence contains all the known canonical MCM motifs including zinc finger, MCM specific Walker A and Walker B and arginine finger. The purified recombinant protein contains ATP-dependent 3′–5′ DNA helicase, ATP-binding and ATPase activities. The helicase activity was stimulated by replication fork like substrate and anti-MCM6 antibodies curtail all the enzyme activities of MCM6 protein. In vitro it self-interacts and forms a homohexamer which is active for DNA helicase and ATPase activities. The complete protein is required for self-interaction as the truncated MCM6 proteins were unable to self-interact. Western blot analysis and in vivo immunostaining followed by confocal microscopy showed the localization of MCM6 both in the nucleus and cytosol. These findings provide first direct evidence that single subunit MCM6 contains DNA helicase activity which is unique to plant MCM6 protein, as this activity was only reported for heteromultimers of MCM proteins in animal system. This discovery should make an important contribution to a better understanding of DNA replication in plants.

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Acknowledgments

Studies on DNA replication and plant abiotic stress tolerance in NT’s laboratory are partially supported by Department of Biotechnology (DBT), Government of India. We thank Dr. Xuan Hoi Pham for his initial help in the manuscript preparation.

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  1. Ngoc Quang Tran

    Present address: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, 240 Longwood Ave, Boston, MA, 02115, USA

Authors and Affiliations

  1. International Centre for Genetic Engineering and Biotechnology, Aruna Asaf Ali Marg, New Delhi, 110067, India

    Ngoc Quang Tran, Hung Quang Dang, Renu Tuteja & Narendra Tuteja

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  1. Ngoc Quang Tran
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  2. Hung Quang Dang
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Correspondence to Narendra Tuteja.

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Tran, N.Q., Dang, H.Q., Tuteja, R. et al. A single subunit MCM6 from pea forms homohexamer and functions as DNA helicase. Plant Mol Biol 74, 327–336 (2010). https://doi.org/10.1007/s11103-010-9675-7

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