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PDBsum entry 4tmc
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PDB id:
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Flavoprotein
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Title:
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Crystal structure of old yellow enzyme from candida macedoniensis aku4588 complexed with p-hydroxybenzaldehyde
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Structure:
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Old yellow enzyme. Chain: a, b, c, d. Engineered: yes
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Source:
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Kluyveromyces marxianus. Yeast. Organism_taxid: 4911. Strain: aku4588. Gene: oye. Expressed in: escherichia coli. Expression_system_taxid: 562
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Resolution:
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1.80Å
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R-factor:
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0.169
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R-free:
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0.205
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Authors:
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S.Horita,M.Kataoka,N.Kitamura,T.Nakagawa,T.Miyakawa,J.Ohtsuka, K.Nagata,S.Shimizu,M.Tanokura
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Key ref:
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S.Horita
et al.
(2015).
An engineered old yellow enzyme that enables efficient synthesis of (4R,6R)-Actinol in a one-pot reduction system.
Chembiochem,
16,
440-445.
PubMed id:
DOI:
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Date:
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31-May-14
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Release date:
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11-Feb-15
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PROCHECK
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Headers
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References
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Q6I7B7
(Q6I7B7_KLUMA) -
Old yellow enzyme from Kluyveromyces marxianus
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Seq:
Struc:
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403 a.a.
399 a.a.
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Key: |
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PfamA domain |
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Secondary structure |
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CATH domain |
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DOI no:
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Chembiochem
16:440-445
(2015)
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PubMed id:
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An engineered old yellow enzyme that enables efficient synthesis of (4R,6R)-Actinol in a one-pot reduction system.
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S.Horita,
M.Kataoka,
N.Kitamura,
T.Nakagawa,
T.Miyakawa,
J.Ohtsuka,
K.Nagata,
S.Shimizu,
M.Tanokura.
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ABSTRACT
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(4R,6R)-Actinol can be stereo-selectively synthesized from ketoisophorone by a
two-step conversion using a mixture of two enzymes: Candida macedoniensis old
yellow enzyme (CmOYE) and Corynebacterium aquaticum (6R)-levodione reductase.
However, (4S)-phorenol, an intermediate, accumulates because of the limited
substrate range of CmOYE. To address this issue, we solved crystal structures of
CmOYE in the presence and absence of a substrate analogue p-HBA, and introduced
point mutations into the substrate-recognition loop. The most effective mutant
(P295G) showed two- and 12-fold higher catalytic activities toward
ketoisophorone and (4S)-phorenol, respectively, than the wild-type, and improved
the yield of the two-step conversion from 67.2 to 90.1%. Our results demonstrate
that the substrate range of an enzyme can be changed by introducing mutation(s)
into a substrate-recognition loop. This method can be applied to the development
of other favorable OYEs with different substrate preferences.
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