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In vitro phosphorylation of the N-terminal half of hordeivirus movement protein

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Abstract

The N-terminal half of TGB1 movement protein of poa semilatent hordeivirus, which forms a ribonucleoprotein complex involved in movement of the viral genome in the plant, and its two domains, NTD and ID, are phosphorylated in vitro by a fraction enriched in cell walls from Nicotiana benthamiana. Using a set of protein kinase inhibitors with different specificities, it was found that enzymes possessing activities of casein kinase 1, protein kinase A, and protein kinase C are involved in phosphorylation. Commercial preparations of protein kinases A and C are able to phosphorylate in vitro recombinant proteins corresponding to the N-terminal half of the protein and its domains NTD and ID. Phosphorylation of the NTD has no effect on the efficiency and character of its binding to RNA. However, phosphorylation of the ID leads to a decrease in its RNA-binding activity and in the ability for homological protein-protein interactions.

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Abbreviations

BaMV:

bamboo mosaic virus

CK:

casein kinase

DTT:

dithiothreitol

ID:

internal domain

IPTG:

isopropyl-β-D-thiogalactopyranoside

MP:

movement protein

NCM:

nitrocellulose membrane

Ni-NTA-agarose:

Ni2+-nitrilotriacetate agarose

N63K:

N-terminal protein half including the NTD and ID

NTD:

N-terminal domain

PK:

protein kinase

PSLV:

poa semilatent virus

PVA:

potato virus A

PVX:

potato virus X

RNP:

ribonucleoprotein

TGB (1–3):

triple gene block encoded proteins (1–3)

TMV:

tobacco mosaic virus

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Authors and Affiliations

  1. Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 119991, Moscow, Russia

    V. V. Makarov, A. Y. Iconnikova & N. O. Kalinina

  2. Biological Faculty, Lomonosov Moscow State University, 119991, Moscow, Russia

    V. V. Makarov & A. Y. Iconnikova

  3. Institute of Agricultural Biotechnology, Russian Academy of Agricultural Sciences, ul. Timiryazevskaya 42, 127550, Moscow, Russia

    M. A. Guseinov & V. K. Vishnichenko

Authors
  1. V. V. Makarov
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  2. A. Y. Iconnikova
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  3. M. A. Guseinov
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  4. V. K. Vishnichenko
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  5. N. O. Kalinina
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Correspondence to N. O. Kalinina.

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Original Russian Text © V. V. Makarov, A. Y. Iconnikova, M. A. Guseinov, V. K. Vishnichenko, N. O. Kalinina, 2012, published in Biokhimiya, 2012, Vol. 77, No. 9, pp. 1292–1302.

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Makarov, V.V., Iconnikova, A.Y., Guseinov, M.A. et al. In vitro phosphorylation of the N-terminal half of hordeivirus movement protein. Biochemistry Moscow 77, 1072–1081 (2012). https://doi.org/10.1134/S0006297912090155

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  • DOI: https://doi.org/10.1134/S0006297912090155

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