Skip to main content

Lund University Publications

LUND UNIVERSITY LIBRARIES

BspK, a serine protease from the predatory bacterium Bdellovibrio bacteriovorus with utility for analysis of therapeutic antibodies

Bratanis, Eleni LU ; Molina, Henrik ; Nägeli, Andreas LU ; Collin, Mattias LU orcid and Lood, Rolf LU (2017) In Applied and Environmental Microbiology 83(4).
Abstract

The development of therapeutic and diagnostic antibodies is a rapidly growing field of research, being the fastest expanding group of products on the pharmaceutical market, and appropriate quality controls are crucial for their application. We have identified and characterized the serine protease termed BspK (Bdellovibrio serine protease K) from Bdellovibrio bacteriovorus and here show its activity on antibodies. Mutation of the serine residue at position 230 rendered the protease inactive. Further investigations of BspK enzymatic characteristics revealed autoproteolytic activity, resulting in numerous cleavage products. Two of the autoproteolytic cleavage sites in the BspK fusion protein were investigated in more detail and... (More)

The development of therapeutic and diagnostic antibodies is a rapidly growing field of research, being the fastest expanding group of products on the pharmaceutical market, and appropriate quality controls are crucial for their application. We have identified and characterized the serine protease termed BspK (Bdellovibrio serine protease K) from Bdellovibrio bacteriovorus and here show its activity on antibodies. Mutation of the serine residue at position 230 rendered the protease inactive. Further investigations of BspK enzymatic characteristics revealed autoproteolytic activity, resulting in numerous cleavage products. Two of the autoproteolytic cleavage sites in the BspK fusion protein were investigated in more detail and corresponded to cleavage after K28 and K210 in the N- and C-terminal parts of BspK, respectively. Further, BspK displayed stable enzymatic activity on IgG within the pH range of 6.0 to 9.5 and was inhibited in the presence of ZnCl2. BspK demonstrated preferential hydrolysis of human IgG1 compared to other immunoglobulins and isotypes, with hydrolysis of the heavy chain at position K226 generating two separate Fab fragments and an intact IgG Fc domain. Finally, we show that BspK preferentially cleaves its substrates C-terminally to lysines similar to the protease LysC. However, BspK displays a unique cleavage profile compared to several currently used proteases on the market.

(Less)
Please use this url to cite or link to this publication:
author
; ; ; and
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
Autoproteolysis, Bdellovibrio bacteriovorus, Biotechnology, Immunoglobulins, Serine protease, Therapeutic antibodies
in
Applied and Environmental Microbiology
volume
83
issue
4
article number
e03037-16
publisher
American Society for Microbiology
external identifiers
  • scopus:85011966183
  • pmid:27940543
  • wos:000393482200021
ISSN
0099-2240
DOI
10.1128/AEM.03037-16
language
English
LU publication?
yes
id
6d25aee6-f14a-4d5d-8b8a-06af69e9c878
date added to LUP
2017-02-23 11:14:18
date last changed
2022-08-02 02:59:53
@article{6d25aee6-f14a-4d5d-8b8a-06af69e9c878,
  abstract     = {{<p>The development of therapeutic and diagnostic antibodies is a rapidly growing field of research, being the fastest expanding group of products on the pharmaceutical market, and appropriate quality controls are crucial for their application. We have identified and characterized the serine protease termed BspK (Bdellovibrio serine protease K) from Bdellovibrio bacteriovorus and here show its activity on antibodies. Mutation of the serine residue at position 230 rendered the protease inactive. Further investigations of BspK enzymatic characteristics revealed autoproteolytic activity, resulting in numerous cleavage products. Two of the autoproteolytic cleavage sites in the BspK fusion protein were investigated in more detail and corresponded to cleavage after K<sub>28</sub> and K<sub>210</sub> in the N- and C-terminal parts of BspK, respectively. Further, BspK displayed stable enzymatic activity on IgG within the pH range of 6.0 to 9.5 and was inhibited in the presence of ZnCl<sub>2</sub>. BspK demonstrated preferential hydrolysis of human IgG1 compared to other immunoglobulins and isotypes, with hydrolysis of the heavy chain at position K<sub>226</sub> generating two separate Fab fragments and an intact IgG Fc domain. Finally, we show that BspK preferentially cleaves its substrates C-terminally to lysines similar to the protease LysC. However, BspK displays a unique cleavage profile compared to several currently used proteases on the market.</p>}},
  author       = {{Bratanis, Eleni and Molina, Henrik and Nägeli, Andreas and Collin, Mattias and Lood, Rolf}},
  issn         = {{0099-2240}},
  keywords     = {{Autoproteolysis; Bdellovibrio bacteriovorus; Biotechnology; Immunoglobulins; Serine protease; Therapeutic antibodies}},
  language     = {{eng}},
  number       = {{4}},
  publisher    = {{American Society for Microbiology}},
  series       = {{Applied and Environmental Microbiology}},
  title        = {{BspK, a serine protease from the predatory bacterium Bdellovibrio bacteriovorus with utility for analysis of therapeutic antibodies}},
  url          = {{http://dx.doi.org/10.1128/AEM.03037-16}},
  doi          = {{10.1128/AEM.03037-16}},
  volume       = {{83}},
  year         = {{2017}},
}