Structural and functional basis for starch binding in the SnRK1 subunits AKIN beta 2 and AKIN beta gamma
Por:
Avila-Castaneda, A, Gutierrez-Granados, N, Ruiz-Gayosso, A, Sosa-Peinado, A, Martinez-Barajas, E, Coello, P
Publicada:
16 may 2014
Categoría:
Plant Science
Resumen:
Specialized carbohydrate-binding domains, the Starch-Binding Domain
(SBD) and the Glycogen Binding Domain (GBD), are motifs of approximately
100 amino acids directly or indirectly associated with starch or
glycogen metabolism. Members of the regulatory beta subunit of the
heterotrimeric complex AMPK/SNF1/SnRK1 contain an SBD or GBD. In
Arabidopsis thaliana, the beta regulatory subunit AKIN beta 2 and a
gamma-type subunit, AKIN beta gamma, also have an SBD. In this work, we
compared the SBD of AKIN beta 2 and AKIN beta gamma with the GBD present
in rat AMPK beta 1 and demonstrated that they conserved the same overall
topology. The majority of the amino acids identified in the
protein-carbohydrate interactions in the rat AMPK beta 1 are conserved
in the two plant proteins. In AKIN beta gamma, there is an insertion of
three amino acids that creates a loop adjacent to one of the conserved
tryptophan residues. Functionally, the SBD from AKIN beta gamma and AKIN
beta 2 could bind starch, but there was an important difference in the
association when an amylose/amylopectin (NA) mixture was used. The
physiological relevance of binding to starch was clear for AKIN beta
gamma, because immunolocalization experiments identified this protein
inside the chloroplast. SnRK1 activity was not affected by the addition
of A/A to the reaction mixture. However, addition of starch inhibited
the activity 85%. Furthermore, proteins associated with A/A and starch
in an in vitro-binding assay accounted for 10-20% of total SnRK1 kinase
activity. Interestingly, the identification of the SnRK1 subunits
associated to the protein-carbohydrate complex indicated that only the
catalytic subunits, AKIN 10 and AKIN11, and the regulatory subunit AKIN
beta gamma were present. These results suggest that a dimer formed
between either catalytic subunit and AKIN beta gamma could be associated
with the A/A mixture in its active form but the same subunits are
inactivated when binding to starch.
Filiaciones:
Univ Nacl Autonoma Mexico, Dept Bioquim, Fac Quim, Mexico City 04510, DF, Mexico
Univ Nacl Autonoma Mexico, Dept Bioquim, Fac Med, Mexico City 04510, DF, Mexico
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