Number of the records: 1

Engineered Lactococcus lactis Secreting IL-23 Receptor-Targeted REX Protein Blockers for Modulation of IL-23/Th17-Mediated Inflammation

SYSNO ASEP	0507592
Document Type	J - Journal Article
R&D Document Type	Journal Article
Subsidiary J	Článek ve WOS
Title	Engineered Lactococcus lactis Secreting IL-23 Receptor-Targeted REX Protein Blockers for Modulation of IL-23/Th17-Mediated Inflammation
Author(s)	Plavec, T. V. (SI) Kuchař, Milan (BTO-N)_RID Benko, A. (SI) Lišková, Veronika (BTO-N) Černý, Jiří (BTO-N)_{RID, ORCID} Berlec, A. (SI) Malý, Petr (BTO-N)_{RID, ORCID}
Number of authors	7
Article number	152
Source Title	Microorganisms. - : MDPI Roč. 7, č. 5 (2019)
Number of pages	13 s.
Language	eng - English
Country	CH - Switzerland
Keywords	lactococcus ; binding protein ; albumin-binding domain ; IL-23 cytokine
Subject RIV	EE - Microbiology, Virology
OECD category	Microbiology
R&D Projects	NV16-27676A GA MZd - Ministry of Health (MZ)
	ED1.1.00/02.0109 GA MŠMT - Ministry of Education, Youth and Sports (MEYS)
Method of publishing	Open access
Institutional support	BTO-N - RVO:86652036
UT WOS	000470962000040
EID SCOPUS	85074238090
DOI	10.3390/microorganisms7050152
Annotation	Lactococcus lactis, a probiotic bacterium of food origin, has recently been demonstrated as a suitable strain for the production and in vivo delivery of therapeutically important proteins into the gut. We aimed to engineer recombinant L. lactis cells producing/secreting REX binding proteins that have been described as IL-23 receptor (IL-23R) blockers and IL-23R antagonists suppressing the secretion of cytokine IL-17A, a pivotal step in the T-helper Th17-mediated pro-inflammatory cascade, as well as in the development of autoimmune diseases, including inflammatory bowel disease (IBD). To reach this goal, we introduced cDNA sequences coding for REX009, REX115, and REX125 proteins into plasmid vectors carrying a Usp45 secretion signal, a FLAG tag sequence consensus, and a LysM-containing cA surface anchor (AcmA), thus allowing cell-surface peptidoglycan anchoring. These plasmids, or their non-FLAG/non-AcmA versions, were introduced into L. lactis host cells, thus generating unique recombinant L. lactis-REX strains. We demonstrate that all three REX proteins are expressed in L. lactis cells and are efficiently displayed on the bacterial surface, as tested by flow cytometry using an anti-FLAG antibody conjugate. Upon 10-fold concentration of the conditioned media, a REX125 secretory variant can be detected by Western blotting. To confirm that the FLAG/non-FLAG REX proteins displayed by L. lactis retain their binding specificity, cell-surface interactions of REX proteins with an IL-23R-IgG chimera were demonstrated by flow cytometry. In addition, statistically significant binding of secreted REX009 and REX115 proteins to bacterially produced, soluble human IL-23R was confirmed by ELISA. We conclude that REX-secreting L. lactis strains were engineered that might serve as IL-23/IL-23R blockers in an experimentally induced mouse model of colitis.
Workplace	Institute of Biotechnology
Contact	Monika Kopřivová, Monika.Koprivova@ibt.cas.cz, Tel.: 325 873 700
Year of Publishing	2020
Electronic address	https://www.mdpi.com/2076-2607/7/5/152

Number of the records: 1