ACTA BIOLOGICA CRACOVIENSIA

BIOSYNTHESIS, GENETICS, AND METABOLISM OF CAROTENOIDS
In biosynthesis of α-carotene in land plants, both lycopene
β-cyclase and lycopene ε-cyclase are needed to produce
α-carotene from lycopene. They have high homology with each
other, and therefore lycopene ε-cyclase gene might be produced
by duplication of lycopene β-cyclase gene. In enzymatic reaction
of cyclization, the mechanisms of lycopene β-cyclase, lycopene
(6'R)-ε-cyclase, and lycopene (6'S)-ε-cyclase are almost the same;
the products are depending on the carbon number to eliminate
H + and on the direction of elimination. Therefore, both lycopene
ε-cyclases could be exist, but only lycopene (6'R)-ε-cyclase was
found based on the presence of only (6'R)-type. Since the stereochemistry
of (6'R)- and (6'S)-α-carotenes are different for the
direction of ε-end groups, the binding site on the protein should
not be identical. Consequently, the protein moiety might restrict
to one chirality of α-carotene, (6'R)-α-carotene.
6.17.
Specific non-natural C50 carotenoid pathways
constructed by the combinatorial expression of
laboratory-evolved enzymes
Maiko Furubayashi 1 , Shinichi Takaichi 2 , Norihiko Misawa 3 ,
Daisuke Umeno 4
1 Department of Applied Chemistry and Biotechnology, Chiba
University, Yayoi, Inage, Chiba 263-8522, Japan,
maiko.furubayashi@gmail.com
2 Department of Biology, Nippon Medical School, Kosugi-cho 2,
Nakahara, Kawasaki 211-0063, Japan, takaichi@nms.ac.jp
3 Research Institute for Bioresources and Biotechnology, Ishikawa
Prefectural University, Suematsu, Nonoichi-machi, Ishikawa
921-8836, Japan, n-misawa@ishikawa-pu.ac.jp
4 Department of Applied Chemistry and Biotechnology, Chiba
University, Yayoi, Inage, Chiba 263-8522, Japan, umeno@faculty.chiba-u.jp
Rapid expansion of available carotenogenic genes has enabled to
produce various carotenoids in heterologous systems. In addition,
non-natural carotenoids can be also produced using combinatorial
expression of laboratory-evolved carotenogenic enzymes
in heterologous hosts of Escherichia coli (Lee et al., 2003; Umeno
et al., 2005). However, carotenogenic enzymes are in general
promiscuous, and it is especially so for the laboratory-evolved
ones. Thus, mere combination of these enzymes results in the
complex mixture of carotenoids containing only a minor amount
of the target. Here, using our C 50 and C 35 carotenogenic pathways
as a model system, we explored how pathway engineers can systematically
'evolve' the specificity of the artificial carotenogenic
pathways. It includes: (1) creation of geranylfarnesyl diphosphate
(C 25 PP) synthase from geranylgeranyl diphosphate (C 20 PP) synthase
CrtE, (2) development of the size mutant of diapophytoene
synthase CrtM, (3) combinatorial expression of above mentioned
mutants for efficient and selective production of C 50 backbone
carotenoids, and (4) desaturation of C 50 carotenoids with the
mutant of phytoene desaturase CrtI. Our current effort to further
diversifying this C 50 pathway, together with extensive analytical
effort, will be also presented.
REFERENCES
LEE PC, MOMEN AZR, MIJTS BN, SCHMIDT-DANNERT C. 2003.
Biosynthesis of structurally novel carotenoids in Escherichia
coli. Chem Biol 10: 453-462.
UMENO D, TOBIAS AV, ARNOLD FH. 2005. Diversifying carotenoid biosynthetic
pathways by directed evolution. Microbiol Mol Biol Rev 69:
51-78.
Vol. 53, suppl. 1, 2011
17–22 July 2011, Krakow, Poland
6.18.
Unique carotenoid lactoside, P457,
in Symbiodinium sp. of dinoflagellate
Takahiro Wakahama1,2 , Hidetoshi Okuyama1,2 ,
Takashi Maoka3 , Shinichi Takaichi4 1Course in Environmental Molecular Biology and Microbial
Ecology, Graduate School of Environmental Science, Hokkaido
University, Sapporo 060-0810, Japan
2Department of Biological Science, School of Science, Hokkaido
University, Sapporo 060-0810, Japan
3Research Institute for Production Development, Sakyo, Kyoto 606-
0805, Japan
4Department of Biology, Nippon Medical School, Kosugi-cho 2,
Nakahara, Kawasaki 211-0063, Japan, takaichi@nms.ac.jp
The dinoflagellates are a large group of unicellular protists common
mostly in marine and also in fresh water environments.
They are characterized by possession of two types of flagellum
with a different direction. About half of dinoflagellates are photosynthetic,
and some of them are known as an endosymbiont of
various kinds of marine animals, such as scleractinian corals, sea
cucumbers, and sea anemones. Symbiotic dinoflagellates are
called zooxanthellae. Another feature of photosynthetic dinoflagellates
is that they have peridinin, a light-harvesting carotenoid
in photosynthesis. In addition to peridinin, an unique carotenoid,
P457, was first descrived by Jeffery in 1968as highlypolar pinkorange
carotenoid from Amphinidium. P457 was also found in
photosynthetic dinoflagellates including zooxanthellae. Its structure
from Amphinidium was determined by Liaaen-Jense group
(1993); neoxanthin-like with an aldehyde group and a lactoside.
Presence of P457 in Symbiodinium derived from marine animals
was not reported. In this study, we reconfirmed the molecular
structure of P457 from Symbiodinium sp. NBRC104787,
isolated from sea anemone, using spectroscopic methods including
CD, FD-MS, and 1H-NMR. In addition, we investigated the distribution
of P457 in various Symbiodinium sp. and scleractinian
coral species, and possible biosynthetic pathways of carotenoids
including P457 are proposed.
A part of this work was supported by Grant-in-Aid for
ScientificResearch on Innovative Areas "Coral reef science for symbiosis
and coexistence of human and ecosystem under combined stresses"
(No.20121002) of the Ministry of Education, Culture, Sports, Science
and Technology (MEXT), Japan.
6.19.
Engineering ketocarotenoid biosynthesis in
maize endosperm
Changfu Zhu1 , Gemma Farre1 , Chao Bai1 , Teresa Capell1 ,
Gerhard Sandmann2 , Paul Christou1,3 1Departament de Producció Vegetal i Ciencia Forestal, Universitat
de Lleida, Av. Alcalde Rovira Roure, 191, Lleida, 25198, Spain,
zhu@pvcf.udl.es, g.farre@pvcf.udl.es, chaobai37@pvcf.udl.es,
capell@pvcf.udl.es
2Biosynthesis Group, Molecular Biosciences, J.W. Goethe
Universitaet, Biocampus 213, P.O. Box 111932, D-60054
Frankfurt, Germany, sandmann@bio.uni-frankfurt.de
3Institucio Catalana de Recerca i Estudis Avancats, Passeig Llúis
Companys, 23, Barcelona 08010, Spain, christou@pvcf.udl.es
Astaxanthin is a highly valued ketocarotenoid with applications in
the nutraceutical, cosmetic, food, and animal feed industries. It is
not a typical plant carotenoid although its precursors β-carotene
and zeaxanthin are abundant. It is possible to genetically engineer
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