BioChem - DOS

Coupled reactions on bioparticles: Stereoselective reduction with cofactor regeneration on PhaC inclusion bodies

Biotechnology journal

Spieler, Valerie; Valldorf, Bernhard; Maaß, Franziska; Kleinschek, Alexander; Hü ttenhain, Stefan H.; Kolmar, Harald

<P><B>Abstract</B></P><P>Chiral alcohols are important building blocks for specialty chemicals and pharmaceuticals. The production of chiral alcohols from ketones can be carried out stereo selectively with alcohol dehydrogenases (ADHs). To establish a process for cost&#8208;effective enzyme immobilization on solid phase for application in ketone reduction, we used an established enzyme pair consisting of ADH from <I>Rhodococcus erythropolis</I> and formate dehydrogenase (FDH) from <I>Candida boidinii</I> for NADH cofactor regeneration and co&#8208;immobilized them on modified poly&#8208;p&#8208;hydroxybutyrate synthase (PhaC)&#8208;inclusion bodies that were recombinantly produced in <I>Escherichia coli</I> cells. After separate production of genetically engineered and recombinantly produced enzymes and particles, cell lysates were combined and enzymes endowed with a Kcoil were captured on the surface of the Ecoil presenting particles due to coiled&#8208;coil interaction. Enzyme&#8208;loaded particles could be easily purified by centrifugation. Total conversion of 4'&#8208;chloroacetophenone to (<I>S</I>)&#8208;4&#8208;chloro&#8208;&alpha;&#8208;methylbenzyl alcohol could be accomplished using enzyme&#8208;loaded particles, catalytic amounts of NAD<SUP>+</SUP> and formate as substrates for FDH. Chiral GC&#8208;MS analysis revealed that immobilized ADH retained enantioselectivity with 99 % enantiomeric excess. In conclusion, this strategy may become a cost&#8208;effective alternative to coupled reactions using purified enzymes.</P>

2016

WILEY&#x2010;VCH Verlag

1860-6768

1860-7314

11

7

pp.890-898

10.1002/biot.201500495

http://click.ndsl.kr/servlet/OpenAPIDetailView?keyValue=05787966&target=NART&cn=NART76491920

Alcohol dehydrogenase; Cofactor regenerating system; Coiled&#x2010; coil interaction; Enzyme immobilization; Multienzyme cascade;