Biocatalyst Engineering: Metabolic Engineering, Kinetic modeling and metagenomics applied to industrial biotechnology

Research output: ThesisDoctoral Thesis (monograph)

Abstract

Industrial biotechnology has been defined as the use and application of
biotechnology for the sustainable processing and production of chemicals,
materials and fuels. It makes use of biocatalysts such as microbial communities,
whole-cell microorganisms or purified enzymes. Although biocatalysts are
considered advantageous, since they operate under mild conditions regarding
temperature and pH and enable chemo-, regio-, and stereoselective reactions,
their utilization on the industrial scale can be impeded by sub-optimal
performance. The present study was aimed at the improvement of two biocatalytic
processes: whole-cell bioreduction for the production of optically pure
alcohols, and ethanol production from lignocellulosic feedstock.
The reduction of the bicyclic diketone, bicyclo[2.2.2]octane-2,6-dione into
1R,4S,6S-6-hydroxy-bicyclo[2.2.2]octane-2-one (endo-alcohol) and 1S,4R,6S-6-
hydroxy-bicyclo[2.2.2]octane-2-one (exo-alcohol) was used as a model bioreduction
reaction. The identification and overexpression of an exo-reductase
encoding gene in Candida tropicalis enabled the production of the uncommon
exo-alcohol as major product. In parallel, the advantages and disadvantages of
metabolically engineered Saccharomyces cerevisiae and Escherichia coli as host
for whole-cell bioreduction were compared for the production of the endoalcohol.
Both these microorganisms gave about the same product purity. While
E. coli showed a three times higher reduction rate, higher cell viability was
maintained during the bioreductions with recombinant S. cerevisiae, which
resulted in higher final conversion (95%) and indicated that yeast could be
recycled.
Improvement of bioethanol production from xylose was achieved through the
construction and use of a kinetic model as a simulation tool for metabolic
engineering of recombinant S. cerevisiae strains. In parallel, novel xylose
isomerases and reductases were isolated from soil metagenome libraries by
sequence- and activity-based screening methods. In addition a novel indirect
protocol for soil DNA extraction that enabled the isolation of environmental
DNA at high yield and purity was developed.
This study enabled the expansion of a biocatalyst toolbox by providing new
catalysts, screening methods and generating new recombinant strains with
improved properties, which can be utilized in the pharmaceutical and bioenergy
sectors, and thus constitutes a step forward in the development of novel biobased
processes.

Details

Authors
  • Nadia Skorupa Parachin
Organisations
Research areas and keywords

Subject classification (UKÄ) – MANDATORY

  • Industrial Biotechnology

Keywords

  • industrial biocatalysts, metagenomics, biocatalyst, Saccharomyces cerevisiae
Original languageEnglish
QualificationDoctor
Awarding Institution
Supervisors/Assistant supervisor
Award date2010 Oct 14
Print ISBNs978-91-7473-011-1
Publication statusPublished - 2010
Publication categoryResearch

Bibliographic note

Defence details Date: 2010-10-14 Time: 10:00 Place: Lecture hall B, Center of Chemistry and Chemical Engineering, Getingevägen 60, Lund University Faculty of Engineering External reviewer(s) Name: Nevoigt, Elke Title: [unknown] Affiliation: Jacobs University, Bremen, Germany ---