Download University of Mitrovica “Isa Boletini”

University of Mitrovica
“Isa Boletini”
Complex media and genetic manipulation for improvement and
production of new product by Kluyveromyces marxianus
Bahtir Hyseni^~ , Rifat Morina^, Alush Musaj^
^Univerity of Mitrovica, Food Technology Department, Faculty of Food Engineering
~Yeditepe University, Graduate School of Natural And Applied Sciences
GREDIT2016 Skopje 01.04.2016
• Outlines
•
Kluyveromyces marxianus
•
Range of metabolites production by K.marxianus in different variety of
medium
•
Ethanol production
•
Other products of K. marxianus
•
Genetic analysis and manipulation of K.marxianus and their expression
•
Conclusion
Role of Kluyveromyces marxianus in
Biotechnology
• Kluyveromyces marxianus is well known yeast, and it
has many known qualities which make it a desirable tool
in biotechnology
• Capabilities, which makes it a desirable organism for
industry like high temperature fermentation and different
kind of medium assimilation.
•
The wide range of the medium assimilation is a suitable
feature for usage a complex media which have cheaper
source.
The main characteristics of K.marxianus
There are many products that are analyzed to be produced
by K.marxianus among them are:
– Ethanol, β-galactosidase , Inulinase
Other characteristics of K. marxianus are;
– high capacity of converting substrate to biomass, growth in high
temperature,
– high production of enzymes,aroma compound production
– good expression of heterologous protein production
– have been labeled as fastest growing eukaryote on the planet
Ethanol production from K.marxianus
The ethanol could be produced from Opuntia ficus-indica and
Jerusalem artichoke juice.
Ethanol production also was found to be successful by using the
juice from Henequen (agave fourcrodyes lem) leaf which are
used as a source of natural fibers.
Mixture of beet molasses and cheese whey was used as substrate
for cultivation of strains of K.marxianus and S.cerevisiae, the
substrate was converted efficiently to ethanol
The results shows a low biomass yield, but the protein content of the
medium was improved.
Different complex medium used for growth
of K.marxianus
Reference Type of medium
Maximum production of Ethanol
g/l
[24]
[20]
K.marxianus
5.6
initial 102
S.cerevisiae
/
/
[21]
D-xylose
Jerusalem artichoke extract with 250 g/l
sugar addition
Jerusalem Artichoke irrigated with 25% sea water
57.5
/
[25]
Mixture of beet molasses and cheese whey
72.6
70.8
[22]
Jerusalem artichoke at High gravity condition with
addition of pectinase and K2HPO4
Opuntia ficus-indica
YDP medium
Henequen juice acid hydrolyzed
Henequen juice enzyme hydrolyzed juice
6.5g/l Apple chokeberry pomace, free cells, without
hydrolysis
6.0g/l Apple cranberry pomace, free cells, without
hydrolysis
6.5g/l Apple chokeberry pomace, immobilized
cells, without hydrolysis
6.0g/l Apple cranberry pomace, immobilized cells,
94.2
/
19.5
69
13.74
16.5
1.37
20.6
/
No growth
No growth
/
1.65
/
1.08
/
0.14
/
[17]
[26]
[27]
[27]
[28]
[28]
[28]
[28]
Different other products of K. marxianus
• β-galactosidase which is produced by K.marxianus
MTCC1388 by using whey.
• It is a great opportunity to use this strain in βgalactosidase production form whey, when is know the
big concern of whey in environment.
• From their results was concluded, that this process can
be applied in industrial scale.
Different other products of K. marxianus
• Inulinase is another product produced by K.marxianus by
using Dahlia tuber extract, which was fortified with yeast
extract as carbon and nitrogen source. The combined medium
shows potential for inulinase production.
• Strains of K.marxianus were successfully used to ferment
Theobroma cacao seeds.
•
Another positive effect was seen in the color of seeds.
Genetic analysis and manipulation of
K.marxianus and their expression
• Although with great ability for many metabolite production
K.marxianus was also subject to many gene transformation
for better expression, adding foreign gene features for
production of new products.
• A good example is development of gene integration method
by non homologous end joining pathway, by using PCRamplified DNA molecules.
• For testing its heterologous expression ability of K.marxianus,
authors Siekstele et. al., analyzed the expression of the EPG
gene (endo-poly-galacturonase) in the yeast by cloning.
Genetic analysis and manipulation of
K.marxianus and their expression
• K.marxianus genes were found to be useful for transformation
of the other species, GAP1 gene which is responsible for
production of the p37 protein, mostly found in the cell of
K.marxianus was found to be doubtful for the flocculation of
the cells.
• Feature of GAP1 gene was proved when non-flocculation
cells
of
Sccharomyces
transformed by GAP1 gene.
cerevisiae
were
genetically
Genetic analysis and manipulation of K.marxianus and their expression
• Another gene of the K.marxianus which was used for cloning
in Sccharomyces cerevisiae was PHO610, which encodes an
acid phosphatase, which is a novel cell-wall protein having an
enzyme activity.
• The high evaluation of the yeast K.marxianus for the ethanol
production was the reason that Nonklang et. al, construct a
flocculation type of the yeast by introducing the gene of
S.cerevisiae FLO1, FLO5, FLO9 and FLO10 by using TDH3
promoter.
Conclusion
• K.marxianus is known yeast recently for its potential to replace some
known “horse work microorganism”.
• High potential for using lignocellulosic and food waste materials by
K.marxianus, makes it known for biotechnology industry.
• The catabolic ability to break down the unused components from
producing process and to turn them in valuable pharmaceutical or
food components is a great strength of these yeasts.
• With improvements in the biotechnology research techniques, many
features of K.marxianus were highlights, with a hope that with a
continuous work in the future this microorganism will be an important
tool for bio-products.