Molecular cloning and expression of human Tumor-associated Polymorphic Epithelial Mucin

/pdf/the-structure-biosynthesis-and-function-of-glycosylated-4rbegebbn2.pdf

The structure, biosynthesis and function of glycosylated phosphatidylinositols in the parasitic protozoa and higher eukaryotes.

The gene for the circumsporozoite (CS) protein of Plasmodium falciparum has been cloned and its nucleotide sequence determined. The gene encodes a protein of 412 amino acids as deduced from the nucleotide sequence. The protein contains 41 tandem repeats of a tetrapeptide, 37 of which are Asn-Ala-Asn-Pro and four of which are Asn-Val-Asp-Pro. Monoclonal antibodies against the CS protein of Plasmodium falciparum were inhibited from binding to the protein by synthetic peptides of the repeat sequence. The CS protein of Plasmodium falciparum and the CS protein of a simian malaria parasite, Plasmodium knowlesi, have two regions of homology, one of which is present on either side of the repeat. One region contains 12 of 13 identical amino acids. Within the nucleotide sequence of this region, 25 of 27 nucleotides are conserved. The conservation of these regions in parasites widely separated in evolution suggests that they may have a function such as binding to liver cells and may represent an invariant target for immunity.

https://science.sciencemag.org/content/sci/225/4662/593.full.pdf

Structure of the Gene Encoding the Immunodominant Surface Antigen on the Sporozoite of the Human Malaria Parasite Plasmodium falciparum

Short-sequence DNA repeat (SSR) loci can be identified in all eukaryotic and many prokaryotic genomes. These loci harbor short or long stretches of repeated nucleotide sequence motifs. DNA sequence motifs in a single locus can be identical and/or heterogeneous. SSRs are encountered in many different branches of the prokaryote kingdom. They are found in genes encoding products as diverse as microbial surface components recognizing adhesive matrix molecules and specific bacterial virulence factors such as lipopolysaccharide-modifying enzymes or adhesins. SSRs enable genetic and consequently phenotypic flexibility. SSRs function at various levels of gene expression regulation. Variations in the number of repeat units per locus or changes in the nature of the individual repeat sequences may result from recombination processes or polymerase inadequacy such as slipped-strand mispairing (SSM), either alone or in combination with DNA repair deficiencies. These rather complex phenomena can occur with relative ease, with SSM approaching a frequency of 10−4 per bacterial cell division and allowing high-frequency genetic switching. Bacteria use this random strategy to adapt their genetic repertoire in response to selective environmental pressure. SSR-mediated variation has important implications for bacterial pathogenesis and evolutionary fitness. Molecular analysis of changes in SSRs allows epidemiological studies on the spread of pathogenic bacteria. The occurrence, evolution and function of SSRs, and the molecular methods used to analyze them are discussed in the context of responsiveness to environmental factors, bacterial pathogenicity, epidemiology, and the availability of full-genome sequences for increasing numbers of microorganisms, especially those that are medically relevant.

https://repub.eur.nl/pub/8835/9618442.pdf

Short-Sequence DNA Repeats in Prokaryotic Genomes

A highly immunogenic region of a human polymorphic epithelial mucin expressed by carcinomas is made up of tandem repeats.

https://www.cell.com/cell/pdf/0092-8674(83)90538-X.pdf

Structure of the plasmodium knowlesi gene coding for the circumsporozoite protein

The malarial sporozoite, the infective stage found in the salivary gland of the insect vector, bears highly immunogenic surface antigen(s). Repeated exposure to irradiated sporozoites induces protection against malaria in several host species, including man. Further, monoclonal antibodies that confer passive immunity react with the immunogenic surface determinants of different sporozoite species. One approach to prevent malaria, therefore, would be to produce a vaccine that induces high titres of circulating antibodies against the sporozoite surface determinant(s). However, production of such a vaccine has not been possible since sporozoites cannot be cultivated in vitro and, therefore, only limited amounts of surface antigen may be obtained. To overcome this problem, we have prepared mRNA from Plasmodium knowlesi-infected mosquitoes to construct a cDNA library. From this library we have isolated a clone that expresses the sporozoite surface antigen as a beta-lactamase fusion protein in the plasmid pBR322. This is the first potentially protective malarial antigen to be cloned by recombinant DNA technology.

Cloning and expression in E. coli of the malarial sporozoite surface antigen gene from Plasmodium knowlesi

Specificity of Tn5 insertions into a 36-bp DNA sequence repeated in tandem seven times

The major surface antigen (circumsporozoite-protein) of the sporozoite stage of Plasmodium knowlesi has been cloned and characterized. The gene is notable for the presence of a 36 base pair unit repeated in tandem 12 times. These repeats may function at the DNA level in regulating gene expression and at the protein level in providing multiple copies of a single epitope (repitope) as part of a protein designed to evade and decoy the immune system.

Structure and Organization of Genes for Sporozoite Surface Antigens

A recombinant complementary DNA clone from Plasmodium knowlesi makes a beta-lactamase fusion polypeptide in Escherichia coli that reacts with a monoclonal antibody to a Plasmodium surface antigen. An epitope of the surface antigen was localized by transposon Tn5 mutagenesis mapping of the complementary DNA clone. The Tn5 mutation having the farthest 5' insert into the complementary DNA portion of the chimeric gene, giving the shortest truncated protein that maintained the ability to bind monoclonal antibody, defined the location of the epitope.

https://science.sciencemag.org/content/sci/220/4603/1285.full.pdf

Luiz S. Ozaki

Papers

Structure of the plasmodium knowlesi gene coding for the circumsporozoite protein

Cloning and expression in E. coli of the malarial sporozoite surface antigen gene from Plasmodium knowlesi

Specificity of Tn5 insertions into a 36-bp DNA sequence repeated in tandem seven times

Structure and Organization of Genes for Sporozoite Surface Antigens

Localization of a Plasmodium surface antigen epitope by Tn5 mutagenesis mapping of a recombinant cDNA clone.