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Thermal-Stable Proteins of Fruit of Long-Living Sacred Lotus Nelumbo nucifera Gaertn var. China Antique

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Abstract

Single-seeded fruit of the sacred lotus Nelumbo nucifera Gaertn var. China Antique from NE China have been shown to remain viable for as long as ~1,300 years, determined by direct radiocarbon-dating, and to have a germination rate of 84 %. The pericarp, a fruit tissue that encloses the single seeds of Nelumbo, is one of the major factors contributing to fruit longevity. Proteins that are heat stable and have a protective function are equally important to such centuries-long seed viability. We document proteins of Nelumbo fruit that are able to withstand heating, 32 % of which remained soluble in the 110 °C-treated embryo axis of a 549-year-old fruit and 76 % retained fluidity in its cotyledons. The genome of Nelumbo has recently been published and annotated. The amino-acid sequences of 11 “thermal proteins” (soluble at 100 °C) of modern Nelumbo embryo axes and cotyledons, identified by mass spectrometry, Western blot and bioassay, are assembled and aligned with those of an archaeal hyperthermophile Methancaldococcus jannaschii (“Mj,” an anaerobic methanogen having a growth optimum of 85 °C) and with those of five mesophile angiosperms. These thermal proteins have roles in protection and repair under stress. More than half (55 %) of the durable Nelumbo thermal proteins are present in the archaean Mj, indicating their ancient history. One Nelumbo protein-repair enzyme exhibits activity at 100 °C, having a heat-tolerance higher than the comparable enzyme of Arabidopsis. A list of 30 sequenced but unassembled thermal proteins of Nelumbo is appended.

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Abbreviations

AdoMet:

S-adenosyl-L-methionine

CPN20/60:

Chaperonine20/60

CuZn-SOD:

Copper-zinc superoxide dismutase

1-CysPRX:

1-Cys peroxiredoxin

ENO1:

Enolase1

EF-1α:

Elongation factor-1α

EST:

Expressed sequence tag

HSP80:

Heat-shock protein 80

LC-MS-MS:

Liquid-chromatography tandem mass-spectrometry

Mj:

Methancaldococcus jannaschii

PIMT:

Protein L-isoaspartyl methyltransferase

ROS:

Reactive oxygen-species

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Acknowledgments

We thank the reviewers for valuable suggestions, C. Haas-Blaby for bioinformatics and protein alignments, J.W. Schopf for constructive comments, J. Lowenson for helpful discussions, and H. Nguyen for compilation assistance of the MS data. For antisera, we are grateful to P. Viitanen for CPN20 and CPN60, T. Close for dehydrin, and B. Downie and D. Martin for PIMT1. Work in SGC laboratory is supported by NIH grant GM206020. We also thank K.O. Stetter whose seminal research on hyperthermophiles stimulated this ‘hot protein’ study of Nelumbo enzymes.

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Authors and Affiliations

  1. IGPP Center for the Study of Evolution and the Origin of Life, Department of Ecology and Evolutionary Biology, University of California, Los Angeles, Geology Building, Room 5676, 595 Charles E. Young Drive East, Los Angeles, CA, 90095-1567, USA

    J. Shen-Miller

  2. Lehrstuhl Mikrobiologie, Regensburg University, Universitat Str. 31, 93053, Regensburg, Germany

    Petra Lindner

  3. Department of Chemistry and Biochemistry, University of California, Los Angeles, 402 Boyer Hall, Hilgard Avenue, Los Angeles, CA, 90095-1569, USA

    Yongming Xie, Kerry Wooding, Rachel R. O. Loo & Joseph A. Loo

  4. Department of Chemistry and Biochemistry, University of California, Los Angeles, 640 Boyer Hall, Hilgard Avenue, Los Angeles, CA, 90095-1570, USA

    Sarah Villa & Steven G. Clarke

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  1. J. Shen-Miller
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Correspondence to J. Shen-Miller.

Additional information

Communicated by Yuval Cohen

J. Shen-Miller for manuscript preparation, overall experimentation and coordination; P. Lindner for initiation of thermal-protein experiments; Y. Xie, K. Wooding, R.R.O. Loo, and J. A. Loo for protein mass-spectrometry; and S. Villa and S. G. Clarke for PIMT experiments.

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Shen-Miller, J., Lindner, P., Xie, Y. et al. Thermal-Stable Proteins of Fruit of Long-Living Sacred Lotus Nelumbo nucifera Gaertn var. China Antique. Tropical Plant Biol. 6, 69–84 (2013). https://doi.org/10.1007/s12042-013-9124-2

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