Novel enzymes for bacterial lignin degradation - University of Warwick

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20 Φεβ 2013 (πριν από 4 χρόνια και 8 μήνες)

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Principal Supervisor
:


Prof T D H Bugg, Department of Chemistry, University of Warwick


Second Supervisor
:


Prof. E Wellington, Department of Life Sciences, University of Warwick


Where will
the student be based
?


The project will be split between Chemistry (enzymology, bioanalytical work) and Life Sciences
(microbiology, molecular biology)


PhD project title:

Novel enzymes for bacterial lignin degradation


Project description:

Lignin is an aromatic polymer found as

15
-
30% of plant cell wall lignocellulose, and is produced as a
by
-
product of cellulosic bioethanol production. It is a potential source of aromatic renewable
chemicals, if bio
-
catalytic methods can be developed for lignin depolymerisation. Previous studie
s on
microbial lignin degradation have focussed on white
-
rot and brown
-
rot fungi, but bacterial lignin
degraders off advantages

for biotechnology for ease of cloning and enzyme overexpression

[1]
.
Using
a UV
-
vis assay for lignin breakdown developed in TDHB
’s group [
2
], we have identified a number of
novel bacterial lignin degraders from environmental samples [
3
], and we have identified a bacterial
lignin peroxidase DypB in
Rhodococcus jostii

RHA1 [
4
]. We are currently using proteomic methods
,

and screening of DNA libraries
,

to identify further novel bacterial lignin
-
degrading enzymes.

The project will involve the cloning and expression of putative lignin
-
degrading enzymes from lignin
-
degrading bacteria.
From our collection of 11 lignin
-
degrad
ing bacteria, candidate genes have been
identified from proteomic data and genome sequencing. Further candidate genes will be identified
from screening of DNA libraries from bacterial lignin degraders, and from metagenomic DNA
libraries. These genes will b
e cloned from the bacterial host, and the
recombinant enzymes will be
expressed in bacterial hosts,
and purified. Pure enzyme will be
assayed against lignin model
compounds, polymeric lignin and lignocellulose
, using assay methods developed in the group [1
-
3]
.
Overexpression of the cloned genes in bacterial hosts will also be investigated as a

strategy for lignin
breakdown, and will be combined with pathway engineering strategies being developed in the group.
High activity enzymes will be tested for product
ion of aromatic chemicals from biomass.



Key
experimental skill
s

involved
:

Molecular cloning of bacterial genes

Molecular microbiology

Protein expression

Enzyme purification

Enzyme assays & kinetics

Bioanalytical methods (HPLC, GC
-
MS, NMR)


References:

1.

“The emerging role for bacteria in lignin degradation and bio
-
product formation” T.D.H. Bugg,
M. Ahmad, E.M. Hardiman, and R. Singh,
Curr. Opin. Biotech
.,
22
, 394
-
400 (2011).

2.

“Development of novel assays for lignin degradation: comparative analysis of bact
erial and
fungal lignin degraders” M. Ahmad, C.R. Taylor, D. Pink, K. Burton, D. Eastwood, G.D. Bending and
T.D.H. Bugg,
Molecular Biosystems
,
6
,815
-
821 (2010).



3.

“Isolation of bacterial strains able to metabolise lignin from screening of environmental sampl
es”

C.R. Taylor, E.M. Hardiman, M. Ahmad, P.D. Sainsbury, P.R. Norris, and T.D.H. Bugg,
J.

Appl.
Microbiol
.,
113
, 521
-
530 (2012).

4.

“Identification of DypB from
Rhodococcus jostii

RHA1 as a lignin peroxidase”
M. Ahmad
, J.N.
Roberts, E.M. Hardiman, R. Singh,
L.D. Eltis, and T.D.H. Bugg,
Biochemistry
,
50
, 5096
-
5107 (2011)



Contact details for application enquiries:


T.D.Bugg@warwick.ac.uk



Keywords:

Lignin degradation

Enzymology

Renewable chemicals production