Next Article in Journal
The Impact of Single- and Double-Strand DNA Breaks in Human Spermatozoa on Assisted Reproduction
Next Article in Special Issue
Scalable High-Performance Production of Recombinant Horseradish Peroxidase from E. coli Inclusion Bodies
Previous Article in Journal
Post-Burn Pruritus
Previous Article in Special Issue
Engineering Biology to Construct Microbial Chassis for the Production of Difficult-to-Express Proteins
Open AccessArticle

Production of Active Recombinant Hyaluronidase Inclusion Bodies from Apis mellifera in E. coli Bl21(DE3) and characterization by FT-IR Spectroscopy

1
FG Environmental Analytics, Process Analytics and Sensors, Institute of Chemical Technology and Analytics, Vienna University of Technology, Getreidemarkt 9/164, 1060 Wien, Austria
2
FG Bioprocess Technology, ICEBE, Vienna University of Technology, Gumpendorferstrasse 1a, 1060 Vienna, Austria
3
FG Integrated Bioprocess Development, ICEBE, Vienna University of Technology, Gumpendorferstrasse 1a, 1060 Vienna, Austria
*
Author to whom correspondence should be addressed.
Int. J. Mol. Sci. 2020, 21(11), 3881; https://doi.org/10.3390/ijms21113881
Received: 26 April 2020 / Revised: 25 May 2020 / Accepted: 27 May 2020 / Published: 29 May 2020
The bacterium E. coli is one of the most important hosts for recombinant protein production. The benefits are high growth rates, inexpensive media, and high protein titers. However, complex proteins with high molecular weight and many disulfide bonds are expressed as inclusion bodies (IBs). In the last decade, the overall perception of these IBs being not functional proteins changed, as enzyme activity was found within IBs. Several applications for direct use of IBs are already reported in literature. While fluorescent proteins or protein tags are used for determination of IB activity to date, direct measurements of IB protein activity are scacre. The expression of recombinant hyaluronidase from Apis mellifera in E. coli BL21(DE3) was analyzed using a face centered design of experiment approach. Hyaluronidase is a hard to express protein and imposes a high metabolic burden to the host. Conditions giving a high specific IB titer were found at 25 °C at low specific substrate uptake rates and induction times of 2 to 4 h. The protein activity of hyaluronidase IBs was verified using (Fourier transform) FT-IR spectroscopy. Degradation of the substrate hyaluronan occurred at increased rates with higher IB concentrations. Active recombinant hyaluronidase IBs can be immediately used for direct degradation of hyaluronan without further down streaming steps. FT-IR spectroscopy was introduced as a method for tracking IB activity and showed differences in degradation behavior of hyaluronan dependent on the applied active IB concentration. View Full-Text
Keywords: Escherichia coli; active inclusion body; recombinant protein production; upstream process development; FT-IR spectroscopy Escherichia coli; active inclusion body; recombinant protein production; upstream process development; FT-IR spectroscopy
Show Figures

Figure 1

MDPI and ACS Style

Schwaighofer, A.; Ablasser, S.; Lux, L.; Kopp, J.; Herwig, C.; Spadiut, O.; Lendl, B.; Slouka, C. Production of Active Recombinant Hyaluronidase Inclusion Bodies from Apis mellifera in E. coli Bl21(DE3) and characterization by FT-IR Spectroscopy. Int. J. Mol. Sci. 2020, 21, 3881.

Show more citation formats Show less citations formats
Note that from the first issue of 2016, MDPI journals use article numbers instead of page numbers. See further details here.

Article Access Map by Country/Region

1
Search more from Scilit
 
Search
Back to TopTop