TY  - JOUR
A1  - Balakrishnan, Karthikeyan
A1  - Andrei-Selmer, Luminita-Cornelia
A1  - Selmer, Thorsten
A1  - Bacher, Michael
A1  - Dodel, Richard
T1  - Comparison of Intravenous Immunoglobulins for Naturally Occurring Autoantibodies against Amyloid-β
JF  - Journal of Alzheimer's Disease
Y1  - 2010
SN  - 1387-2877
VL  - 20
IS  - 1
SP  - 135
EP  - 143
ER  - 
TY  - CHAP
A1  - Muffler, Kai
A1  - Tippkötter, Nils
A1  - Ulber, Roland
ED  - Timmis, Kenneth N.
T1  - Chemical feedstocks and fine chemicals from other substrates
T2  - Handbook of hydrocarbon and lipid microbiology. Volume 4:  Consequences of microbial interactions with hydrocarbons, oils and lipids. - (Springer reference)
Y1  - 2010
SN  - 978-3-540-77588-1
U6  - http://dx.doi.org/10.1007%2F978-3-540-77587-4_214
SP  - 2891
EP  - 2902
PB  - Springer
CY  - Berlin [u.a.]
ER  - 
TY  - JOUR
A1  - Ribitsch, D.
A1  - Karl, W.
A1  - Birner-Gruenberger, R.
A1  - Gruber, K.
A1  - Eiteljoerg, I.
A1  - Remler, P.
A1  - Wieland, S.
A1  - Siegert, Petra
A1  - Maurer, Karl-Heinz
A1  - Schwab, H.
T1  - C-terminal truncation of a metagenome-derived detergent protease for effective expression in E. coli
JF  - Journal of biotechnology
N2  - Recently, a new alkaline protease named HP70 showing highest homology to extracellular serine proteases of Stenotrophomonas maltophilia and Xanthomonas campestris was found in the course of a metagenome screening for detergent proteases (Niehaus et al., submitted for publication). Attempts to efficiently express the enzyme in common expression hosts had failed. This study reports on the realization of overexpression in Escherichia coli after structural modification of HP70. Modelling of HP70 resulted in a two-domain structure, comprising the catalytic domain and a C-terminal domain which includes about 100 amino acids. On the basis of the modelled structure the enzyme was truncated by deletion of most of the C-terminal domain yielding HP70-C477.

This structural modification allowed effective expression of active enzyme using E. coli BL21-Gold as the host. Specific activity of HP70-C477 determined with suc-l-Ala-l-Ala-l-Pro-l-Phe-p-nitroanilide as the substrate was 30 ± 5 U/mg compared to 8 ± 1 U/mg of the native enzyme. HP70-C477 was most active at 40 °C and pH 7–11; these conditions are prerequisite for a potential application as detergent enzyme. Determination of kinetic parameters at 40 °C and pH = 9.5 resulted in KM = 0.23 ± 0.01 mM and kcat = 167.5 ± 3.6 s⁻¹. MS-analysis of peptide fragments obtained from incubation of HP70 and HP70-C477 with insulin B indicated that the C-terminal domain influences the cleavage preferences of the enzyme. Washing experiments confirmed the high potential of HP70-C477 as detergent protease.
Y1  - 2010
U6  - http://dx.doi.org/10.1016/j.jbiotec.2010.09.947
SN  - 1873-4863 (E-Journal); 0168-1656 (Print)
VL  - 150
IS  - 3
SP  - 408
EP  - 416
PB  - Elsevier
CY  - Amsterdam
ER  -