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Cell-free synthetic biology as key enabling technology for novel food ingredients Marc Struhalla, CEO c-LEcta GmbH BIO World Congress of Industrial Biotechnology Montréal, 24.07.2017
Cell-free synthetic biology as key enabling
technology for novel food ingredients
Marc Struhalla, CEO c-LEcta GmbH
BIO World Congress of Industrial Biotechnology
Montréal, 24.07.2017
Company introduction
At a glance
c-LEcta is one of the world’s leading industrial
biotechnology company located in Leipzig,
Germany
Our core technological competence is the
engineering of enzymes and microorganisms
We provide biotechnological process
innovation for valuable products for the food,
feed and pharma industry
24.07.2017
Company Introduction
│3
BioCube Leipzig
BIO-WCIB 2017 c-LEcta.com
Staff of 50+ highly qualified people.
Profitable operational business since 2014
State-of-the-art research & production facility
with about 2.300 m2 including a pilot-scale
production and scale-up infrastructure.
Established premium technology partner and
supplier of biotech products to the industry
Company Introduction
│4
At a glance
Fermentation suite c-LEcta
BIO-WCIB 2017 c-LEcta.com 24.07.2017
Company introduction
│5
c-LEcta’s technology platform
BIO-WCIB 2017 c-LEcta.com
Strain Engineering
Biodiversity
New enzymes and
microbial strains from
natural sources
Enzyme Engineering
Optimized enzymes
obtained by directed
evolution
Efficient microbial
productions strains
obtained by metabolic
engineering
Optimized production
processes for ingredients
and enzymes
Bioprocess
Development
24.07.2017
│6
Company introduction
2 Product categories
Industrial
enzymes
…produced by enzymatic
synthesis or fermentation
+ Ingredients
& additives
For applications in pharma, feed & food
…used as processing aids
in industrial production
BIO-WCIB 2017 c-LEcta.com 24.07.2017
│7
c-LEcta products in the market
Company introduction
BIO-WCIB 2017 c-LEcta.com
Acrylaway HighT
CalB immo Plus
DENARASE
Food Enzyme; Asparaginase. Reduces
acrylamide formation if food products
Immobilized lipase. Used in pharma &
in the processing of lipids and oils
Nuclease. Used in the production of
vaccines & biopharmaceuticals
BIOCATALYSTSCustomized enzymes for the synthesis
of chemical drugs
distribution partners
Undisclosed customers
24.07.2017
Cell-free, synthetic biology
Cell-free synthetic biology
│9
Cell-free synthetic biology
BIO-WCIB 2017 www.c-LEcta.com
Use of artificial, engineered enzymes for one-pot,
multi-enzyme biotransformations
Main contra arguments: Conversion costs are two high. Enzymes make
use of very costly cofactors or cosubstrates and are too expensive
Product formation rate per time with the enzyme
amount obtained from one liter of fermentation broth:
Product titers obtainable in efficient enzymatic
synthesis processes:
Enzyme cost contribution per kilo of product under
recycling of process engineered enzymes:
> 1,000 g/h/l
> 400 g/l
< 0.1 EUR / kg
24.07.2017
│10
Two examples from c-LEcta work
BIO-WCIB 2017 www.c-LEcta.com
Cost impact of enzymes
120-fold
reduction
Cost impact of cofactor NAD+
25-fold
reduction
24.07.2017
Cell-free synthetic biology
Improvement of process economics through
enzyme engineering & process development
Enzyme cascade for glucosylation
│11
Sucrose-
Synthase
Glucosyl-
Transferase
UDP-Glucose
TARGET
COMPOUND
TARGET
COMPOUND
UDP
Fructose Sucrose
BIO-WCIB 2017 www.c-LEcta.com 24.07.2017
Cell-free synthetic biology
Enzyme cascade for glucosylation
│12
TARGET
COMPOUND
TARGET
COMPOUND
Sucrose
+ +
Net Reaction:
Targets for an efficient process:
High substrate & product concentration
As low as possible UDP concentration
Highly efficient enzymes (low enzyme costs)
Fructose
Targets for enzyme engineering:
Reduce substrate & product inhibition
Decrease Km-values for UDP
Increase specific activity of enzymes
BIO-WCIB 2017 www.c-LEcta.com 24.07.2017
Cell-free synthetic biology
UDP
Enzyme cascade for glucosylation
│13
UDP cost contribution:
Non-optimized system:
[UDP] 0.25 mM
[Product] 25 g/L
Price UDP 2,300 € / kg
Cost contribution UDP:
10.0 € / kg Product
BIO-WCIB 2017 www.c-LEcta.com 24.07.2017
Cell-free synthetic biology
Optimized system:
[UDP] 0.05 mM
[Product] 150 g/L
Price UDP 2,300 € / kg
Cost contribution UDP:
0.34 € / kg Product
Engineering of a Glucosyl-Transferase
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0
5
10
15
20
25
30
35
40
45
50
fold
im
pro
vem
ent
Variants
WT
4-mutations
5-mutations
BIO-WCIB 2017 www.c-LEcta.com 24.07.2017
Cell-free synthetic biology
Sugar phosphorylase cascade
│15
Sugar-
Phosphorylase
Sucrose-
Phosphorylase
(SP)
Glucose-1-
Phosphate
TARGET
DISACCHARIDE
(Glc-Glc)
Fruktose
Glucose
Sucrose
+
Glucose-
Isomerase
Pi +
BIO-WCIB 2017 www.c-LEcta.com 24.07.2017
Cell-free synthetic biology
DISACCHARIDE
PRODUCTNet Reaction:
Sucrose
Pi
0
0.02
0.04
0.06
0.08
0.1
250 500 750
init
ial a
cti
vit
iy[s
yn
the
sis
un
its
]
substrate concentration [mM]
Engineering of a cellobiose phosphorylase (CP)
Below
detection limit
G1P cellobiose + PiCP
glucose
| 16BIO-WCIB 2017 www.c-LEcta.com 24.07.2017
Cell-free synthetic biology
0
5
10
15
20
25
30
35
fold
im
pro
ve
me
nt
250 mM substrate
500 mM substrate
750 mM substrate
Cellobiose phosphorylase engineering
1st gen to
stabilize backbone
WO2016038141
| 17BIO-WCIB 2017 www.c-LEcta.com 24.07.2017
Cell-free synthetic biology
0
5
10
15
20
25
30
35
fold
im
pro
ve
me
nt
250 mM substrate
500 mM substrate
750 mM substrate
1st gen to
stabilize backbone
WO2016038141
| 18
Cellobiose phosphorylase engineering
BIO-WCIB 2017 www.c-LEcta.com 24.07.2017
Cell-free synthetic biology
Very low enzyme cost contribution
Process is economically viable
Synthesis process has been established & scaled-up by partner
Contact details
Marc StruhallaCEO
c-LEcta GmbH
Perlickstraße 5, 04103 Leipzig, Germany
Phone +49 341 / 355 214-0
www.c-LEcta.com
