Enzymes . Chemicals . Solutions .

www.enzymicals.com

NEWPROT Kick-off meeting: 25.01.2012

Partner profile Enzymicals AGDr. Rainer Wardenga

Start-up history

Spin off from the Institute of Biochemistry at the University of Greifswald

Founded in August 2009 as corporation

Management

Dr. Ulf Menyes (CEO) Dr. Rainer Wardenga (CSO)

Supervisory Board

Prof. Uwe Bornscheuer (Chairman)Dr. Dagmar BraunDr. Jürgen Eck

Ownership structure

Shareholders and strategic investor

1

Northern Germany, Biotechnikum Greifswald

Company Info

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1

Company Info

www.enzymicals.com

Northern Germany, Sinetific Campus Greifswald

Start-up history

Spin off from the Institute of Biochemistry at the University of Greifswald

Founded in August 2009 as corporation

Management

Dr. Ulf Menyes (CEO) Dr. Rainer Wardenga (CSO)

Supervisory Board

Prof. Uwe Bornscheuer (Chairman)Dr. Dagmar BraunDr. Jürgen Eck

Ownership structure

Shareholders and strategic investor

Management and staff

Dr. U. Menyes (Chemist), CEO

Dr. R. Wardenga (Biologist and Biotechnologist), CSO Dipl. S. Weber (Economist), finance and controlling

Biotechnologists (main focus molecular biology)

Biotechnologist (main focus biocatalysis)

Chemical technical assistant

Biological technical assistant

2

Company Info

www.enzymicals.com

Scientific staff

Company management

Business Segments

3

Company Info

www.enzymicals.com

Institute of Biochemistry at the University of Greifswald (Prof. Bornscheuer)

B.R.A.I.N. AG

HERBRAND PharmaChemicals GmbH

Institute of Chemistry at the University of Rostock (Prof. Kragl)

Institute of Marine Biotechnology e.V. Greifswald

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4

Partners

Research & Development Along the Value Chain

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5

Core competence in protein science and development of biocatalytic process

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6

Research & Development Along the Value Chain

Identification of suitable catalysts

Method development and routine application of HPLC, CE, GC and GC-MS

Design and application of spectrophotometric assaysSynthesis of screening substrates

Screening of enzymes or strain collections Search in literature or pathway databasesSearch of known or novel motives in sequence

databases

Amplification of novel genes from genomic DNADe-novo genome sequencing and analysis

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Cloning, expression and purification

Fully equipped molecular laboratoriesState of the art expertise in molecular cloningHuge repertoire of expression vectors

Well-founded knowledge of a range of recombinant expression systems

Approved solutions for proteins which are difficult to express

Screening of refolding conditions for aggregated proteins

Production capacity up to 30 L-scaleCatalysts are available in different grades of purity

Purification via affinity-chromatography or fusion-tags

Research & Development Along the Value Chain

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Cloning, expression and purification in collaboration with B.R.A.I.N AG

Peptide and Protein Production in

· Escherichia coli / Bacillus sp.

· Pseudomonas stutzeri / Pichia pastoris

· Streptomyces sp.

· Aspergillus sp.

Production Systems:

Optimized Production of Peptides or Proteins in Industrial Scale

Research & Development Along the Value Chain

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Biochemical, physiochemical and structural characterization

Determination of catalytic properties and kinetic parameters

Characteristics in temperature- and pH-profilesInvestigation of storage and process stability

Properties via gel-electrophoresis and blottingSecondary structure via CD-spectrophotometry

Screening for crystallization conditions via commercial kits

Access to MALDI-TOF, 2D-gel-electrophoresis

General immobilization studies

Research & Development Along the Value Chain

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Protein engineering

Selection of best engineering approaches for designated targets

Approaches often inspired by structural alignments or molecular modeling

Homology-model construction and hot-spot identification Rational design of variants by site-directed mutagenesis

Directed evolution with high quality variant librariesSelection assays and library screening with high-

throughput systems

HT-screening of mutant-librariesAccess to 3-laser flow cytometer with cell sorter

Research & Development Along the Value Chain

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Application of best catalysts paired with optimized chemistry

Innovative synthetic routes using organic synthesis and biotransformation

Custom manufacturing of fine and special chemicals up to the multi-kg scale

Catalysis in multi-phase systems or ionic liquids Membrane processes for catalyst recovery and

downstream processing

Research & Development Along the Value Chain

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Application of best catalysts paired with optimized chemistry

Innovative synthetic routes using organic synthesis and biotransformation

Custom manufacturing of fine and special chemicals up to the multi-kg scale

Catalysis in multi-phase systems or ionic liquids Membrane processes for catalyst recovery and

downstream processing

Scope of our biocatalytic tollbox

Research & Development Along the Value Chain

www.enzymicals.com

13

NEWPROT: Objectives

(R)-selective amine-transaminases

Catalyze the transfer an amino group from a primary amine to a prochiral ketone or aldehyde

Usable for the resolution of racemic amines or the synthesis of chiral amines

Pyridoxal-5‘-phosphate dependent

Typical amines : alanine, ethylamine, 1- and 2-propylamine, 1- and 2-butylamine ….

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(R)-selective amine-transaminases

Catalytic tool box for the evaluation of synthesis routes

Conversion of aliphatic, arylaliphatic and aromatic ketones with strict (R)-selectivity

pH-optima range between 7.0 and 8.5

Alanine serves as amine donor

Products obtained by asymmetric synthesis with ECS-ATA01-07.

NEWPROT: Objectives

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Baeyer-Villiger monooxygenases

Flavoenzymes from the class of oxidoreductases

Oxidation of linear, cyclic and aromatic ketones to esters or lactones

Usable with cofactor regeneration or as whole cell system

Known applications:

Biooxidation of functionalized cycloketones

Biooxidation of bicyclic ketones or chiral b-substituted cycloketones

Kinetic resolution of racemic a-substituted cycloketones

Regiodivergent biooxidation of fused bicyclobutanones

NEWPROT: Objectives

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Baeyer-Villiger monooxygenases

Catalytic tool box for the evaluation of synthesis routes

Baeyer-Villiger oxidation of aromatic and aliphatic ketones, able to perform sulfur oxidations

High regio- and enantioselectivity, NAD(P)H-dependent

Scope of the ECS biooxidation-toolbox. Examples of products obtained from the corresponding ketones with ECS-M01-07.

O

HN O

O

ee > 99%

OO

OO

MeMe

ee > 99%

ee > 99%

S

O

O

O

Me

ee > 98%

O

H

H

O

ee > 99%

O

ee > 98%

O

Cl

O

HN O

O

ee > 99%

OO

OO

MeMe

ee > 99%

ee > 99%

S

O

O

O

Me

ee > 98%

O

H

H

O

ee > 99%

O

ee > 98%

O

Cl

NEWPROT: Objectives

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Thank you for your attention

Enzymicals AG,

Walther-Rathenau-Straße 49a17489 GreifswaldFon: +49(0)3834 515-47-0Fax: +49(0)3834 515-47-3