Cloning of usp45, a gene encoding a secreted protein from Lactococcus lactis subsp. lactis MG1363

Asseldonk, Martien van; Rutten, Ger; Oteman, M.; Siezen, Roland J.; Vos, Willem M. de; Simons, Guus

doi:10.1016/0378-1119(90)90428-t

Cited by 203 publications

(135 citation statements)

References 17 publications

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“…This vector contains the following features ( Fig. 2): the lactococcal pNZ212 origin of replication; a chloramphenicol acetyltransferase gene for selection; the P nisA promoter to direct inducible expression of PA fusions [23]; the lactococcal USP45 secretion signal to drive secretion [24]; the gene fragment encoding the PA (the C-terminal 218 amino acid cell-wall binding domain of AcmA of L. lactis [25] including its stop codon); and a transcriptional terminator is present 445 bp downstream of the PA gene fragment. Genes can be inserted in the unique NcoI site, resulting in a transcriptional fusion with the Met residue located at position +5 of the mature secreted protein.…”

Section: Antigen-protein Anchor Fusionsmentioning

confidence: 99%

Mucosal vaccine delivery of antigens tightly bound to an adjuvant particle made from food-grade bacteria

Roosmalen¹,

Kanninga²,

Khattabi³

et al. 2006

Methods

106

102

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Section: Antigen-protein Anchor Fusionsmentioning

confidence: 99%

Mucosal vaccine delivery of antigens tightly bound to an adjuvant particle made from food-grade bacteria

Roosmalen¹,

Kanninga²,

Khattabi³

et al. 2006

Methods

106

102

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“…The final construct was named pSEC-huLTN. This strategy permitted the fusion in-frame of the DNA sequence coding for LTN with the signal peptide from the usp45 (SP Usp45 ), the most secreted protein of L. lactis (van Asseldonk et al 1990), and placed under the regulation of the well-characterized nisin-inducible promoter (PnisA) (Fig. 1).…”

Section: Resultsmentioning

confidence: 99%

Production of biologically active human lymphotactin (XCL1) by Lactococcus lactis

et al. 2008

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Lymphotactin-XCL1 is a chemokine produced mainly by activated CD8? T-cells and directs migration of CD4? and CD8? lymphocytes and natural killer (NK) cells. We expressed human lymphotactin (LTN) by the lactic-acid bacterium Lactococcus lactis. Biological activity of LTN was confirmed by chemo-attraction of human T-cells by chemotaxis demonstrating, for the first time, how this chemokine secreted by a food-grade prokaryote retains biological activity and chemoattracts T lymphocytes. This strain thus represents a feasible well-tolerated vector to deliver active LTN at a mucosal level.

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“…These plasmids are derived from two broad-host-range expression vectors, pCYT:Nuc and pSEC:Nuc (Bermudez-Humaran et al, 2003). The former contains i) the PxylT promoter (Jamet, 2001), ii) the ribosome-binding site of the lactococcal usp45 gene (van Asseldonk et al, 1990), and iii) the DNA fragment encoding the mature part of the staphylococcal nuclease, NucB (Le Loir et al, 1994). The latter contains all of this DNA sequence, plus the signal peptide of the Usp45 coding sequence (spUsp45), necessary for the secretion of NucB.…”

Section: Construction Of Recombinant L Lactis Strains To Produce Eitmentioning

confidence: 99%

Cytoplasmic and extracellular expression of pharmaceutical-grade mycobacterial 65-kDa heat shock protein in Lactococcus lactis

Azevedo

Rocha²,

Electo³

et al. 2012

Genet. Mol. Res.

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ABSTRACT. Lactic acid bacteria (LAB) are an attractive and safe alternative for the expression of heterologous proteins, as they are nonpathogenic and endotoxin-free organisms. Lactococcus lactis, the LAB model organism, has been extensively employed in the biotechnology field for large-scale production of heterologous proteins, and its use as a "cell factory" has been widely studied. We have been particularly interested in the use of L. lactis for production of heat shock proteins (HSPs), which reportedly play important roles in the initiation of innate and adaptive immune responses. However, this activity has been questioned, as LPS contamination appears to be responsible for most, if not all, immunostimulatory activity of HSPs. In order to study the effect of pure HSPs on the immune system, we constructed recombinant L. lactis strains able to produce and properly address the Mycobacterium leprae 65-kDa HSP (Hsp65) to the cytoplasm or to the extracellular medium, using a xylose-induced expression system. Expression of mycobacterial Hsp65 in Lactococcus lactisApproximately 7 mg/L recombinant Hsp65 was secreted. Degradation products related to lactococcal HtrA activity were not observed, and the Limulus amebocyte lysate assay demonstrated that the amount of LPS in the recombinant Hsp65 preparations was 10-100 times lower than the permitted levels established by the U.S. Food and Drug Administration. These new L. lactis strains will allow investigation of the effects of M. leprae Hsp65 without the interference of LPS; consequently, they have potential for a variety of biotechnological, medical and therapeutic applications.

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Cloning of usp45, a gene encoding a secreted protein from Lactococcus lactis subsp. lactis MG1363

Cited by 203 publications

References 17 publications

Mucosal vaccine delivery of antigens tightly bound to an adjuvant particle made from food-grade bacteria

Mucosal vaccine delivery of antigens tightly bound to an adjuvant particle made from food-grade bacteria

Production of biologically active human lymphotactin (XCL1) by Lactococcus lactis

Cytoplasmic and extracellular expression of pharmaceutical-grade mycobacterial 65-kDa heat shock protein in Lactococcus lactis

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