%0 Journal Article
%A Christenson, James
K.
%A Robinson, Serina L.
%A Engel, Tiffany A.
%A Richman, Jack E.
%A Kim, An N.
%A Wackett, Larry P.
%D 2017
%T OleB from Bacterial Hydrocarbon Biosynthesis Is a
β‑Lactone Decarboxylase That Shares Key Features with
Haloalkane Dehalogenases
%U https://acs.figshare.com/articles/journal_contribution/OleB_from_Bacterial_Hydrocarbon_Biosynthesis_Is_a_Lactone_Decarboxylase_That_Shares_Key_Features_with_Haloalkane_Dehalogenases/5421670
%R 10.1021/acs.biochem.7b00667.s001
%2 https://acs.figshare.com/ndownloader/files/9353398
%K Protein sequence analyses
%K Gram-negative bacterium Xanthomonas campestris
%K alkylated wild-type X
%K nucleophilic aspartic acid
%K oleABCD gene clusters
%K OleB D 114A
%K Bacterial Hydrocarbon Biosynthesis
%K trans -β- lactones
%K Shares Key Features
%K OleB proteins
%K HLD-III
%K Gram-positive bacterium Micrococcus luteus
%K h 2 18 O studies
%K biosynthesize long-chain olefinic hydrocarbons
%K β- lactone decarboxylation reaction
%K decarboxylating cis -β- lactones
%K β- lactone decarboxylation
%K HLD
%K Haloalkane Dehalogenases OleB
%K β- lactone decarboxylases
%X OleB
is an α/β-hydrolase found in bacteria that biosynthesize
long-chain olefinic hydrocarbons, but its function has remained obscure.
We report that OleB from the Gram-negative bacterium Xanthomonas
campestris performs an unprecedented β-lactone decarboxylation
reaction, to complete cis-olefin biosynthesis. OleB
reactions monitored by 1H nuclear magnetic resonance spectroscopy
revealed a selectivity for decarboxylating cis-β-lactones
and no discernible activity with trans-β-lactones,
consistent with the known configuration of pathway intermediates.
Protein sequence analyses showed OleB proteins were most related to
haloalkane dehalogenases (HLDs) and retained the canonical Asp-His-Asp
catalytic triad of HLDs. Unexpectedly, it was determined that an understudied
subfamily, denoted as HLD-III, is comprised mostly of OleB proteins
encoded within oleABCD gene clusters, suggesting
a misannotation. OleB from X. campestris showed very
low dehalogenase activity only against haloalkane substrates with
long alkyl chains. A haloalkane substrate mimic alkylated wild-type X. campestris OleB but not OleBD114A, implicating
this residue as the active site nucleophile as in HLDs. A sequence-divergent
OleB, found as part of a natural OleBC fusion and classified as an
HLD-III, from the Gram-positive bacterium Micrococcus luteus was demonstrated to have the same activity, stereochemical preference,
and dependence on the proposed Asp nucleophile. H218O studies with M. luteus OleBC suggested
that the canonical alkyl–enzyme intermediate of HLDs is hydrolyzed
differently by OleB enzymes, as 18O is not incorporated
into the nucleophilic aspartic acid. This work defines a previously
unrecognized reaction in nature, functionally identifies some HLD-III
enzymes as β-lactone decarboxylases, and posits an enzymatic
mechanism of β-lactone decarboxylation.
%I ACS Publications