Research Overview
Research group for microbial carbohydrate metabolic engineering
Introduction
Research group for microbial carbohydrate metabolic engineering is engaged in
the development of commercially important microorganisms by means of metabolic
engineering.
At present, the main research program is focused on the improvement
of yeast strains, which can effectively ferment both glucose and xylose from
lignocellulosic biomass to produce ethanol.
Also, the group has engineered other
yeast strains to produce pyruvate and xylitol respectively. Pyruvate and xylitol are
the important bio-products and intermediates of microbial carbohydrate metabolism.
Professor Ning Jiang, head of the group, graduated from the Graduate School of
Chinese Academy of Sciences in 1981. He has worked in the fields of applied
microbiology and microbial biotechnology for more than twenty years in the Institute
of Microbiology, CAS.
Since the mid 1990s, his research interest has focused on
metabolic engineering to improve industrial microbial strains.
He is the member of
the board of the Chinese Journal of Biotechnology and the Chinese Journal of Applied
and Environmental Biology.
The group has four other staffs and seven doctorial students.
The staffs are An
Shen (associate professor), Peng He (research associate), Shuhao Wang (research
associate) and Dajun Lu (technician). The students are Changying Guo, Hua Yan,
Changsheng Wu, Hairong Chen, Pingying Wei, Zilong Li and Yuping Lin.
Background and Significance
Yeast, especially Saccharomyces cerevisiae is the main material for the research
of the group. Saccharomyces cerevisiae has a lot of advantages such as well studied
genetic background, high fermentation rates with the ability to grow under both
aerobic and anaerobic conditions, ethanol tolerance and osmo-tolerance, easy to
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manipulate and safety for foods.
Fuel ethanol as an alternative energy source is becoming more and more
important triggered by the world energy crisis.
Lignocellulosic biomass composed
of cellulose, hemicellulose and lignin is the most abundant and cheap bio-resource on
the earth.
Using lignocelluloses as raw material to produce ethanol faces great
opportunities and challenges.
Saccharomyces cerevisiae is an ideal ethanol producer
but can’t ferment xylose, the main degradation product of hemicellulose. One of the
key problems for producing ethanol from lignocellulosic biomass is to make the
microorganism xylose fermentable.
Many efforts of the group are to introduce
xylose metabolic pathway and set up the balance of co-factors in Saccharomyces
cerevisiae cells.
Major Achievements
The group is presently working on the National High-tech R&D Program of
China titled “research of recombinant strains for fuel ethanol and efficient cellulase”,
the National Basic Research Program of China titled “basic research of key processes
for applications of lignocellulosic biomass” and the National Knowledge Innovation
Project of CAS titled “breeding and optimization of microorganism for effective
conversion of biomass”.
The major achievements of the group in the past four years
were:
1, Construction of novel Saccharomyces cerevisiae strain which can co-ferment
glucose and xylose to produce ethanol effectively
Xylose reductase gene (XYL1), xylitol dehydrogenase gene (XYL2) and xylulose
kinase gene (XKS1) from various yeasts were expressed in Saccharomyces cerevisiae.
Also, bacterial transhydrogenase gene (udhA) from E. coli was cloned and expressed
in the recombinant yeast.
The novel engineered strain could ferment xylose and
regenerate NADPH simultaneously so that it could effectively ferment both glucose
and xylose to produce ethanol under oxygen-limited condition.
The conversion
yields of glucose and xylose to ethanol were 90% and 85% respectively.
2, Improvement of various producing behaviors of Saccharomyces cerevisiae for
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ethanol production
Various producing behaviors of Saccharomyces cerevisiae for ethanol production
were improved by mutagenesis and DNA recombination.
These behaviors included
conversion yield, productivity and rate of ethanol fermentation, osmo-tolerance,
ethanol tolerance and thermo-tolerance.
In laboratory experiments, the strains
obtained by the group could ferment glucose or sucrose to produce ethanol with the
yield of 95%, the productivity of 18 % (v/v) and the rate of 3 g/l.h. The yeast could
grow and ferment in the medium containing 300 g/l glucose and tolerant 20% ethanol.
Also, 10 % (v/v) of ethanol was produced when fermented at 42 C.
3, Construction of pyruvate hyper-producing strain
The pdc gene encoding pyruvate decarboxylase in Torulopsis glabrata was
specifically disrupted. The disruptant displayed higher pyruvate accumulation and less
ethanol production.
It accumulated 80 g/l of pyruvate in 100 L jar pilot fermentation
under favorable conditions.
4, Construction of xylitol producing strain
The xylose reductase gene from Pichia stipitis was cloned and expressed in
Saccharomyces cerevisiae. The recombinant yeast could convert xylose to xylitol
with the yield of 90 %.
5, Publications
P He, D-J Lu, A Shen, N Jiang* :Cloning and expression of bacterial hemoglobin
gene in D-arabitol producing yeast, Acta Microbiol. Sinica, 41（3）：315－319，2001
Q-H Wang, P He, D-J Lu, A Shen, N Jiang* :Screening of pyruvate-producing
yeast and effect of nutritional conditions on pyruvate production ， Lett. Appl.
Microbiol., 35: 338 – 342, 2002
Q-H Wang, P He, D-J Lu, A Shen, N Jiang* : Purification, Characterization,
Cloning and Expression of Pyruvate Decarboxylase from Torulopsis glabrata IFO005,
J. Biochem., 136: 447 - 455, 2004
X-L Liu, P He, D-J Lu, A Shen N, Jiang*:Construction of flocculation selective
vector and expression of β-glucosidase gene in Saccharomyces cerevisiae，Chin. J.
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Biotechnol., 21(1)：167-170,2005
Q-H Wang, P He, D-J Lu, A Shen, N Jiang* :Metabolic Engineering of
Torulopsis glabrata for Improved Pyruvate Production，Enzyme Microb. Technol.，
36:832-839,2005
X-L Liu, N Jiang*, P He, D-J Lu, A Shen :Fermentation of xylose to produce
ethanol by recombinant Saccharomyces cerevisiae strain containing XYLA and XKS1,
Chinese Sci. Bull.,50:652-657,2005
H-R Cheng ,N Jiang,* A Shen, Y-J Feng: Molecular cloning and functional
expression of D-arabitol dehydrogenase gene from Gluconobacter oxydans in
Escherichia coli, submitted to FEMS Microbiol Lett.
H-R Cheng and N Jiang* : Extremely rapid and efficient extraction of DNA from
bacteria and yeasts using a novel method, submitted to Biotechnol. Lett.
C-Y Guo, P He, D Lu, A Shen, N Jiang* : Screening and characterization of
xylose-utilizing microorganism for xylitol production, submitted to Lett Appl
Microbiol.
C-Y Guo, P He, D Lu, A Shen, N Jiang* :Cloning, molecular characterization
and enzymatic analysis of CsXYL3 (D-Xylulokinase) from Candida sp. Xu316 and its
expression in Saccharomyces cerevisiae submitted to J. Appl. Microbiol.
C-Y Guo, P He, D Lu, A Shen, N Jiang* : Redox cofactor regeneration in
recombinant Saccharomyces cerevisiae to improve ethanol production from xylose
submitted to Meta. Engin.
6, Patents:
N. Jiang et al: A novel method for producing xylitol by cell conversion,
ZL96120000.6 (2001)
N. Jiang et al: A novel method for producing D-arabitol from glucose by yeast
cells, ZL99119504.3 (2002)
N. Jiang et al: Xylitol production by multiple reused free yeast cells,
ZL011110778.2 (2004)
N. Jiang et al: Pyruvate production by fermentation 02122657.8
N. Jiang et al: A recombinant yeast strain for pyruvate production, 03109214.4
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N. Jiang et al: Pyruvate production method and its specific strain,
200410033847.6
N. Jiang et al: Xylitol production by nixed fermentation, 200410031949.4
N. Jiang et al: Xylitol production method and its specific strain, 200510008756.1
Future Research Plan
The main research interest of the group in the near future will still focus on fuel
ethanol according to the urgent request for energy source in China. Also, pyruvate
and xylitol producing strains will be commercialized.
1, Development of economically feasible processes for ethanol production from
lignocellulosic biomass
The hexose-pentose co-fermentation yeast strains will be further improved.
The group will try to solve the problem of how to degrade lignocellulose effectively,
economically and environment friendly.
It is the most important problem for using
lignocellulosic materials to produce ethanol.
2, Improvement of industrial ethanol producing yeast
Various ethanol producing yeast strains that possess different favorable
characteristics have been obtained.
The further effort is to integrate multiple
favorable characteristics in one strain by genome shuffling and high-throughput
screening. The novel strain will be more useful for industrial ethanol production.
3, Commercialization of pyruvate and xylitol
Pyruvate and xylitol producing strains will be further improved and applied in
commercial production by co-operation with companies.
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