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Identical active sites in hydroxynitrile lyases show opposite enantioselectivity and reveal possible ancestral mechanism.


ABSTRACT: Evolutionarily related hydroxynitrile lyases from rubber tree (HbHNL) and from Arabidopsis thaliana (AtHNL) follow different catalytic mechanisms with opposite enantioselectivity toward mandelonitrile. We hypothesized that the HbHNL-like mechanism evolved from an enzyme with an AtHNL-like mechanism. We created ancestor-like composite active-sites in each scaffold to elucidate how this transition may have occurred. Surprisingly, a composite active site in HbHNL maintained (S)-selectivity, while the identical set of active site residues in AtHNL maintained (R)-selectivity. Composite active-site mutants that are (S)-selective without the Lys236 and Thr11 that are required for the classical (S)-HNL mechanism suggests a new mechanism. Modeling suggested a possibility for this new mechanism that does not exist in modern enzymes. Thus, the last common ancestor of HbHNL and AtHNL may have used an extinct mechanism, not the AtHNL-like mechanism. Multiple mechanisms are possible with the same catalytic residues and residues outside the active site strongly influence mechanism and enantioselectivity.

SUBMITTER: Jones BJ 

PROVIDER: S-EPMC5546752 | biostudies-literature | 2017 Jun

REPOSITORIES: biostudies-literature

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Identical active sites in hydroxynitrile lyases show opposite enantioselectivity and reveal possible ancestral mechanism.

Jones Bryan J BJ   Bata Zsófia Z   Kazlauskas Romas J RJ  

ACS catalysis 20170515 6


Evolutionarily related hydroxynitrile lyases from rubber tree (<i>Hb</i>HNL) and from <i>Arabidopsis thaliana</i> (<i>At</i>HNL) follow different catalytic mechanisms with opposite enantioselectivity toward mandelonitrile. We hypothesized that the <i>Hb</i>HNL-like mechanism evolved from an enzyme with an <i>At</i>HNL-like mechanism. We created ancestor-like composite active-sites in each scaffold to elucidate how this transition may have occurred. Surprisingly, a composite active site in <i>Hb<  ...[more]

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