TITLE:
Further Insights on the Datura innoxia Hyoscyamine 6β-Hydroxylase (DiH6H) Based on Biochemical Characterization and Molecular Modeling
AUTHORS:
Daniel Schlesinger, Faris Salama, Rachel Davidovich Rikanati, Rotem Sertchook, Ofir Tal, Mosaab Yahyaa, Adi Faigenboim, Mwafaq Ibdah, Moshe Inbar, Efraim Lewinsohn
KEYWORDS:
Hyoscyamine 6β Hydroxylase, Datura innoxia. Mill., Solanaceae, Scopolamine, Hyoscyamine
JOURNAL NAME:
American Journal of Plant Sciences,
Vol.12 No.1,
January
22,
2021
ABSTRACT: Hyoscyamine, anisodamine
and scopolamine are tropane alkaloids present
in some Solanaceae species and used in modern medicine. L-Hyoscyamine is hydroxylated
to 6β-hydroxyhyoscyamine
(anisodamine) and then epoxidated to scopolamine
by the dual action of hyoscyamine 6β-hydroxylase
(H6H), a 2-oxoglutarate dependent dioxygenase.
A natural mutation in the Gly-220 residue to Cys was previously shown to be associated
with the loss of function of H6H in Mandragora officinarum, preventing
the accumulation of anisodamine and scopolamine in these plants. We show here
that a deliberate Gly220Cys mutation in the Datura innoxia DiH6H protein caused a loss of both its enzymatic abilities
and rendered it unable to hydroxylate L-hyoscyamine into anisodamine and to epoxidate anisodamine into scopolamine. By using protein modeling
based on an available crystal structure of H6H from Datura metel, we show how the Cys220 residue causes
a steric interference in the active site cavity impairing the interaction of both
substrates, hyoscyamine and anisodamine with the active site of the protein. We also address the enantiomeric
preference of DiH6H based on molecular modeling.