Biomedical subjects

Jos F M L Seegers

Publications and source records attributed to Jos F M L Seegers.

6 recordsLinked to original sources

PubMed2006

Lactobacilli-expressed single-chain variable fragment (scFv) specific for intercellular adhesion molecule 1 (ICAM-1) blocks cell-associated HIV-1 transmission across a cervical epithelial monolayer.

The vaginal and cervical epithelia provide an initial barrier to sexually acquired HIV-1 infection in women. To study the interactions between HIV-1-infected cells or cell-free HIV-1 and the reproductive epithelium, the transmission of HIV-1 by infected cells or cell-free virus across human cervical epithelial cells was examined using a Transwell culture system. Cell-associated HIV-1 was transmitted more efficiently than cell-free virus, and monocyte-associated virus was transmitted most efficiently. Abs to ICAM-1 added to the apical side of the epithelium blocked cell-mediated transepithelial HIV-1 transmission in vitro. When used in a previously described model of vaginal HIV-1 transmission in human PBL-SCID mice, anti-murine ICAM-1 Abs (0.4 microg/10 microl) also blocked vaginal transmission of cell-associated HIV-1 in vivo. To evaluate a candidate delivery system for the use of this Ab as an anti-HIV-1 microbicide, anti-ICAM single-chain variable fragment Abs secreted by transformed lactobacilli were evaluated for their protective efficacy in the Transwell model. Like the intact Ab and Fab derived from it, the single-chain variable fragment at a concentration of 6.7 microg/100 microl was able to reduce HIV-1 transmission by 70 +/- 5%. These data support the potential efficacy of an anti-ICAM Ab delivered by lactobacilli for use as an anti-HIV-1 microbicide.

Animals↗

PubMed2006

Anatomy of a lactococcal phage tail.

Bacteriophages of the Siphoviridae family utilize a long noncontractile tail to recognize, adsorb to, and inject DNA into their bacterial host. The tail anatomy of the archetypal Siphoviridae lambda has been well studied, in contrast to phages infecting gram-positive bacteria. This report outlines a detailed anatomical description of a typical member of the Siphoviridae infecting a gram-positive bacterium. The tail superstructure of the lactococcal phage Tuc2009 was investigated using N-terminal protein sequencing, Western blotting, and immunogold transmission electron microscopy, allowing a tangible path to be followed from gene sequence through encoded protein to specific architectural structures on the Tuc2009 virion. This phage displays a striking parity with lambda with respect to tail structure, which reenforced a model proposed for Tuc2009 tail architecture. Furthermore, comparisons with lambda and other lactococcal phages allowed the specification of a number of genetic submodules likely to encode specific tail structures.

Electrophoresis, Polyacrylamide Gel↗

PubMed2004

Molecular and transcriptional analysis of the temperate lactococcal bacteriophage Tuc2009.

The genome of bacteriophage Tuc2009 consists of 38347 base pairs on which 57 open reading frames (ORFs) were identified, divided in two oppositely transcribed regions. The leftward-transcribed region harbors three ORFs, two of which are involved in the establishment of lysogeny. The rightward-transcribed region contains 54 ORFs, which are assumed to be required for the lytic life cycle. An exception to the above organization is ORF 10, of unknown function, located within the rightward-transcribed region that has an orientation opposite to the ORFs surrounding it. Transcriptional analysis of the Tuc2009 genome following infection of a sensitive host revealed that most ORFs are transcribed in a sequential manner. ORFs that are presumed to form (part of) the genetic switch along with the superinfection exclusion-encoding gene are transcribed immediately after infection, followed by transcription of the presumed replication region. Subsequent to this, several small transcripts could be identified followed by a single 24-kb transcript. This latter transcript was shown to specify most of the identified structural proteins as well as two proteins required for host lysis. Interestingly, the 24-kb mRNA was shown to undergo splicing through the activity of a type I intron whose removal from the mRNA resulted in the formation of an ORF specifying a major structural protein. Primer extension analysis was employed to identify the 5' ends of mRNA transcripts and the genome and transcriptional data are discussed in relation to other lactococcal bacteriophages.

Bacteriophages↗

PubMed2002

Lactobacilli as live vaccine delivery vectors: progress and prospects.

Evidence is accumulating that lactobacilli influence the immune response in a strain-dependent manner. This immunomodulatory capacity is important for the development of the immune response, and also identifies Lactobacillus as a potent oral vaccine carrier. Most of our current knowledge of the use of lactobacilli for vaccination purposes has been obtained with tetanus toxin fragment C (TTFC) as the model antigen. This knowledge, together with our ever-increasing understanding of the immune system and recent developments in cloning and expression techniques, should enable the utilisation of antigens other than TTFC and has made the development of lactobacilli as live vaccines a realistic prospect.

Bacterial Vaccines↗

PubMed2000

Molecular characterization of the lactococcal plasmid pCIS3: natural stacking of specificity subunits of a type I restriction/modification system in a single lactococcal strain.

A 6.1 kb plasmid from the Lactococcus lactis subsp. cremoris strain UC509.9, named pCIS3, was found to mediate a restriction/modification (R/M) phenotype. Nucleotide sequence analysis of pCIS3 revealed the presence of an hsdS gene, typical of type I R/M systems. The presence of this plasmid resulted in a 10(4)-fold reduction in the efficiency of plating (e.o.p.) of unmodified phage. In addition to the hsdS gene of pCIS3, two more hsdS genes were identified in strain UC509.9, one located on the chromosome downstream of a gene highly homologous to hsdM genes and a third on the smallest (4 kb) plasmid, named pCIS1. The replication region of pCIS3 was highly similar to that of a large family of lactococcal theta replicons. In addition, pCIS3 was found to encode a member of the CorA family of magnesium transporters.

Amino Acid Sequence↗

PubMed1997

Purification of two Bacillus subtilis proteins which cross-react with antibodies directed against eukaryotic protein kinase C, the His HPr kinase and trigger factor.

As in eukaryotes, phosphorylation of Ser residues in proteins appears to be common phenomenon in bacteria. Surprisingly, however, very few Ser/Thr protein kinases have been identified and in this study antibodies directed against mammalian protein kinase C (PKC) have been used in attempts to isolate conserved Ser/Thr protein kinases. Using the mAb M7 against rat brain PKC, a single 70 kDa band was identified in total cell extracts of Bacillus subtilis by Western blotting after SDS-PAGE, whilst using polyclonal antibody alpha-PKC1p against Saccharomyces cerevisiae PKC a single 67 kDa band was identified by the same procedure. The two proteins were purified independently on the basis of antibody recognition employing two-dimensional gel electrophoresis as a final step, which allowed subsequent microsequencing. The 70 kDa band was thus identified as the phosphoenolpyruvate-dependent His HPr kinase, Enzyme 1 of the phosphotransferase system. This identity was confirmed using a mutant deleted for ptsl, encoding Enzyme 1. The 67 kDa protein was identified as a previously unknown B. subtilis 'trigger factor', homologous to an Escherichia coli protein-folding enzyme, peptidylprolyl cis-trans-isomerase implicated in cell division.

Amino Acid Isomerases↗