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The plastidial starch phosphorylase from rice endosperm: catalytic properties at low temperature

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Abstract

Main conclusion

Consistent with its essential role in starch biosynthesis at low temperatures, the plastidial starch phosphorylase from rice endosperm is highly active at low temperature. Moreover, contrary to results on other higher plant phosphorylases, the L80 peptide, a domain unique to plant phosphorylases and not present in orthologous phosphorylases from other organisms, is not involved in enzyme catalysis.

Starch phosphorylase (Pho) is an essential enzyme in starch synthesis in develo** rice endosperm as the enzyme plays a critical role in both the early and maturation phases of starch granule formation especially at low temperature. In this study, we demonstrated that the rice Pho1 maintains substantial enzyme activity at low temperature (<20 °C) and its substrate affinities for branched α-glucans and glucose-1-phosphate were significantly increased at the lower reaction temperatures. Under sub-saturating substrate conditions, OsPho1 displayed higher catalytic activities at 18 °C than at optimal 36 °C, supporting the prominent role of the enzyme in starch synthesis at low temperature. Removal of the highly charged 80-amino acid sequence L80 peptide, a region found exclusively in the plastidial Pho1 of higher plants, did not significantly alter the catalytic and regulatory properties of OsPho1 but did affect heat stability. Our kinetic results support the low temperature biosynthetic role of OsPho1 in rice endosperm and indicate that its L80 region is unlikely to have a direct enzymatic role but provides stability of the enzyme under heat stress.

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Abbreviations

Glc1P:

Glucose-1-phosphate

Glc1,6P:

Glucose-1,6-diphosphate

Glc6P:

Glucose-6-phosphate

IPTG:

Isopropyl β-D-1-thiogalactopyranoside

Pi:

Inorganic phosphate

SDS:

Sodium dodecyl sulfate

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Acknowledgments

This work was supported by the grants from the Department of Biotechnology, Government of India as DBT Overseas Associateship (S.S.), from the Japan Society for the Promotion of Science (H.S.), from the Division of Chemical Sciences, Geosciences, and Biosciences, Office of Basic Energy Sciences of the US Department of Energy (Grant DE-FG02-12ER20216 to T.W.O, S.K.H., and B.C.) and from Project 0590, Agricultural Research Center, College of Agricultural, Human, and Natural Resource Sciences, Washington State University.

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    Authors and Affiliations

    1. Institute of Biological Chemistry, Washington State University, Pullman, WA, 99164, USA

      Seon-Kap Hwang, Bilal Cakir & Thomas W. Okita

    2. Department of Agricultural Biotechnology, Assam Agricultural University, Jorhat, Assam, 785013, India

      Salvinder Singh

    3. Faculty of Agriculture, Kyushu University, Fukuoka, 812-8581, Japan

      Hikaru Satoh

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    1. Seon-Kap Hwang
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    2. Salvinder Singh
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    Correspondence to Thomas W. Okita.

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    Seon-Kap Hwang and Salvinder Singh have contributed equally to this work.

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    Hwang, SK., Singh, S., Cakir, B. et al. The plastidial starch phosphorylase from rice endosperm: catalytic properties at low temperature. Planta 243, 999–1009 (2016). https://doi.org/10.1007/s00425-015-2461-7

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