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The evolutionary appearance of non‐cyanogenic hydroxynitrile glucosides in the L otus genus is accompanied by the substrate specialization of paralogous β–glucosidases resulting from a crucial amino acid substitution
Author(s) -
Lai Daniela,
Abou Hachem Maher,
Robson Fran,
Olsen Carl Erik,
Wang Trevor L.,
Møller Birger L.,
Takos Adam M.,
Rook Fred
Publication year - 2014
Publication title -
the plant journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.058
H-Index - 269
eISSN - 1365-313X
pISSN - 0960-7412
DOI - 10.1111/tpj.12561
Subject(s) - lotus corniculatus , biochemistry , lotus japonicus , amino acid , biology , stereochemistry , biosynthesis , chemistry , enzyme , botany , mutant , gene
Summary L otus japonicus , like several other legumes, biosynthesizes the cyanogenic α–hydroxynitrile glucosides lotaustralin and linamarin. Upon tissue disruption these compounds are hydrolysed by a specific β–glucosidase, resulting in the release of hydrogen cyanide. L otus japonicus also produces the non‐cyanogenic γ‐ and β–hydroxynitrile glucosides rhodiocyanoside A and D using a biosynthetic pathway that branches off from lotaustralin biosynthesis. We previously established that BGD 2 is the only β–glucosidase responsible for cyanogenesis in leaves. Here we show that the paralogous BGD 4 has the dominant physiological role in rhodiocyanoside degradation. Structural modelling, site‐directed mutagenesis and activity assays establish that a glycine residue (G211) in the aglycone binding site of BGD 2 is essential for its ability to hydrolyse the endogenous cyanogenic glucosides. The corresponding valine (V211) in BGD 4 narrows the active site pocket, resulting in the exclusion of non‐flat substrates such as lotaustralin and linamarin, but not of the more planar rhodiocyanosides. Rhodiocyanosides and the BGD 4 gene only occur in L . japonicus and a few closely related species associated with the L otus corniculatus clade within the L otus genus. This suggests the evolutionary scenario that substrate specialization for rhodiocyanosides evolved from a promiscuous activity of a progenitor cyanogenic β–glucosidase, resembling BGD 2, and required no more than a single amino acid substitution.