SABMiller has grown into a global company with operations and distribution agreements across six continents.

It is one of the worlds top 3 largest brewers, and its portfolio of brands Pilsner Urquell, Peroni Nastro Azzurro, Castle, Miller Genuine Draft and Grolsch. Local beverage market is saturated, and therefore are interested in some diversification into biotech, especially around byproducts.

Tongaat Hulett Starch is Africas largest producer of Starch, Glucose and related products. It uses enzymes for production of glucose and fructose syrup. Hulett Sugar has a central refinery in South Africa with an annual refining capacity of some 600 000 tons with refining capacity at other sites. Also provides industrial sweeteners Acesulfame K, Aspartame, Sodium Cyclamate Sodium Saccharin, Polydextrose Illovo: Sugar and by products: Furfural, Furfuryl Alcohol, Diacetyl, 2,3 Pentanedione, Methanol, Ethyl alcohol (Ethanol). TSB: Sugar SMRI: Joint research institute, seeking new biotechnology options, particularly natural polymers.

AECI (SA Bioproducts) 11 000 tonne/year lysine plant at Umbogintwini, Natal province, South Africa.

Ciba-BASF nitrile hydratase based biocatalysis technology has been implemented by SENMIN in South Africa.

Senmins new plant, Sasolburg, generates acrylamide from acrylonitrile

UK and SA. BBI Enzymes is the market leader in the manufacture of natural enzymes to a global marketplace.

Natural Enzymes

SA SME start up

Importer

Fermentation for Ethanol Spirit vinegar Fusel oils Interested in Oxidation of alkanes or alkenes and Carbon dioxide use. Environmental biotechnology. Chem City research and SME incubator.

Fragrances, flavours and oils. Products and research include application of biocatalysis (Steenkamp et al (patent)).

Vaccines under development include a liquid pentavalent vaccine (diphtheria, tetanus, pertussis, hepatitis B and Haemophilus influenza type b.

EPO, Monoclonal Antibodies

Streptomyces antibiotics: Actinomycin D ; Brefeldin A ; Ionophores; Leptomycin B etc.

 Bioprospecting: Natural Products Chemistry Emerging Health Technologies (including synthetic biology) Protein Technologies:
Vaccines Antibodies Enzymes Biocatalysis

 Sunlight and dry areas useful for open ponds. eta-carotene production using algal technology. Natural Carotenoids South Africa (Pty) Ltd small plant built in Uppington.

Commercial scale Custom production of plant made proteins

 For a developing country, South Africa has an unusually large chemical industry and it is of substantial economic significance. In 2010 the industry still contributes 5% to the GDP and 25% of manufacturing sales. The chemical industry is focusing on the need to be internationally competitive and the industry is reshaping itself accordingly. Exports were R49 billion and imports R95 billion in 2009 with the gap declining. Local Giants SASOL, AECI, Sentrachem (DOW), OMNIA (Protea) (and about 1000 small companies). Many multi-national companies operate in South Africa as manufacturers and/or distributors. Amongst these companies are Air Products, Bayer, BASF, Buckman, du Pont, Huntsman Tioxide, Lanxess, Merck and Rohm and Haas (now part of Dow). (Source CAIA 2010)

 Environmentally Friendly Technology Aim to enhance the fine chemicals and Pharmaceutical sectors. Interest in renewables

Hydrogenation of thymol produces an 8 isomer mixture of the enantiomers of menthol, isomenthol, neomenthol and neoisomenthol). CSIR patented process utilises an enzyme in organic media to enantioselectively and diastereoselectively catalyse the separation of ()-menthol from the post hydrogenation mixture

CSIR 2011 www.csir.co.za

OH O N N

N O
+

O NH N H O

NH NH2

HO

OH

Guanosine

Thymine

PO43-

Purine nucleoside phosphorylase Uridine nucleoside phosphorylase O O NH N

+

2 step 2 enzyme 1 pot high yielding biocatalytic reaction in a slurry. 20% solid loading ~90% yield Integrated into synthesis of thymidine, stavudine and AZT
O NH NH OH O MeOH, NaOMe, 40C, 1 h HO N O O

PNP from halophile

NH N N H2

OH O

N H

Directed evolution of UP from E. coli

Visser DF, Hennessy F, Rashamuse K, Louw M and Brady D. (2010) Cloning, purification and characterisation of the purine nucleoside phosphorylase from Bacillus halodurans Alk36. Extremophiles 14: 185-192 Visser DF, Rashamuse JK, Hennessy F, Gordon G, van Zyl PJ, Mathiba K, Bode ML and D Brady (2010). Cascade Enzymatic Synthesis of 5-Methyluridine Using a Novel Combination of Nucleotide Phosphorylases. Biocatalysis and Biotransformation 28: 245-253. Gordon GER, Visser DF, Bode ML, Raseroka N.and Brady D (2011). A high yielding multienzyme biocatalytic reaction for the synthesis of 5methyluridine by transglycosylation of guanosine and thymine. Journal of Biotechnology, 151: 108-113. Gordon GER, Visser DF, Bode M L, Lepuru J., Zeevaart JG, Ragubeer N, Ratsaka M, Walwyn D and D Brady (2011). A chemo-enzymatic route for preparation of -thymidine, a key intermediate in the preparation of antiretrovirals. Organic Process Research and Development 15: 258-265. Visser DF, Hennessy F, Rashamuse J, Pletschke B and Brady D (2011). Stabilization of Escherichia coli uridine phosphorylase by evolution and immobilization. J. Mol. Cat. B: Enz, 68: 279-285.

OH HO 5-Methyluridine AcBr CH3CN, reflux 1 h O NH OAc O N O

Guanine

OAc O

OAc DABT

Br

MeOH, H2 9 bar, Ni catalyst 1-2 h

OAc DAT

Thymidine

CSIR 2011 www.csir.co.za

MeO 2C NHCbz MeO 2 C NHCbz O N H RR/ SS-9 O HO 2C NHCbz O N H SS-12 O a N H O R R-9 O

RR-Monatin is 2000 fold sweeter than sucrose.

Enzymatic resolution of RR/SS monatin derivatives. Reagents and Conditions: a) ChiroCLEC-BL, ACN, phosphate buffer (0.3 M, pH 7.5) 22C, 15-20 h.
MeO 2 C NHCbz O N H RR-9 O N H 14 a HO2 C NHCbz OH CO 2H N H 2 b HO 2C NH2 OH CO2 H

Reagents and Conditions: a) RR-9 in DCM, aq. KOH, 25 C, 30 min, then AcOH; b) 10% Pd/C (5%), MeOH, Et3N, H2 (3 bar), 40 C, 1 h.

CSIR 2012 www.csir.co.za

Rousseau AL, Buddoo SR, Gordon GER, Beemadu S, Kupi BG, Lepuru MJ, Maumela MC, Parsoo A, Sibiya DM, and Brady D (2011). A chemo-enzymatic process for stereoselective synthesis of monatin. Organic Process Research and Development 15: 249-257.

R1 R3

R2

Epoxide hydrolase H2O

R1 R3

R2

OH

Starting epoxide

Blockbuster 2-benzylmorpholine (appetite suppressant)

H N O

* * R 4
Racemic epoxide

+
R4 Optically active chiral epoxide

R1 R3 OH

R2
(2,3 -Epoxypropyl)benzene

R4

Optically active chiral diol

Styrene oxide
O

(S) -Fluoxetine (Prozac)

(anti-depressant)
C F3

O N H

p - Nitrostyrene oxide
O NO 2

(R) Nifnalol (beta blocker)

OH N H iP r NO 2

Chiral Epoxide use in Pharmaceuticals

Cardiovascular agents beta-blockers Antitussive drugs Antibiotics Antiviral agents MAO inhibitors Biochemical probes

trans -1-Phenylpropene oxide

O

Ephedrine (stimulant)
OH NH C H3

Indene oxide**
O

Indinavir (Crixivan) HIV protease inhibitor

C 6H 5 C H2 O N N N C O N H C ( C H 3) 3 OH H N OH

CSIR 2012 www.csir.co.za

DEPA NH2 HO O Amidase HO OH O

HO O KCN

N Nitrile hydratase

Phenylacetaldehyde

2-Hydroxy-3phenyl-propionitrile

2-Hydroxy-3phenyl-propionamide

2-Hydroxy-3-phenylpropionic acid

Cascade reaction: -hydroxynitriles from aldehydes

Strand RF Strand RF Strand RF plus 1 plus 1 plus 1 plus 2 plus 2 plus 2 plus 3 plus 3 plus 3 minus 1 minus 1 minus 1 minus 2 minus 2 minus 2 minus 3 minus 3 minus 3 1 1 1 198 198 198 397 397 397 595 595 595

cDNA LNHase beta alpha chap Universal code cDNA LNHase beta alpha chap cDNA LNHase beta alpha chap Universal code 794 Universal code1191 993 1390 794 993 1191 1390 794 993 1191 1390

1588 1588 1588

1787 1787 1787

1986 1986 1986

Chaperone
Beta Alpha

CSIR 2012 www.csir.co.za

Brady D, Beeton A, Kgaje C, Zeevaart J, van Rantwijk F, Sheldon RA (2004) Characterisation of Nitrilase and Nitrile Hydratase Biocatalytic Systems. Applied Microbiology and Biotechnology. 64: 76-85.

Cross linked porous polyethyleneimine (PEI) matrix Binding of functional molecules throughout the volume of microspheres. PEI has high functional group density. These features translate to exceptional binding capacity, and enable microsphere properties to be readily engineered for performance superiority.

CSIR 2011

www.csir.co.za

Globally Sub-Saharan Africa is estimated as being one of the two regions with the highest biomass (Smeets, 2004).

SA has large sugar industry and Pulp and paper industry.

TIA project (jointly with Universities) and COST action

Biorefining and Carbon cycling program, University of Georgia (http://www.biorefinery.uga.edu/)

Industrial Biotechnology (general) Bio-based Economy (BE-BASIC in NL) Fermentation (Strong in NL) Biocatalysis (Strong in NL) Functional Protein Production Enzymes Bioremediation