Biomedical subjects

S Raffioni

Publications and source records attributed to S Raffioni.

24 records · Page 2Linked to original sources

PubMed1988

Primary structure of the mating pheromone Er-1 of the ciliate Euplotes raikovi.

The complete amino acid sequence of the mating pheromone Er-1 purified from Euplotes raikovi homozygous for mat-1 was determined by automated Edman degradation of the whole protein and peptides generated by cyanogen bromide, trypsin, Staphylococcus aureus V8 protease, and chymotrypsin. The proposed sequence is: Asp-Ala-Cys-Glu-Gln-Ala-Ala-Ile-Gln-Cys-Val-Glu-Ser-Ala-Cys-Glu-Ser-Leu- Cys-Thr-Glu-Gly-Glu-Asp-Arg-Thr-Gly-Cys-Tyr-Met-Tyr-Ile-Tyr-Ser-Asn-Cys- Pro-Pro-Tyr-Val The calculated molecular weight is 4411.0, which is in agreement with the averaged mass of 4410.2 obtained by fission fragment ionization mass spectrometry. Previously reported values of the native molecular weight, determined by gel filtration, have ranged from 9,000 to 12,000. Thus, the native structure is likely a dimer (or larger aggregate) of identical subunits with the three disulfide bonds present occurring as intrachain links. Secondary structure predictions suggest a helical structure at the amino terminus. A comparison of the Er-1 amino acid sequence with known protein sequences did not reveal any significant similarities.

Amino Acid Sequence↗

PubMed1987

Purification and characterization of new mating pheromones of the ciliate Euplotes raikovi.

Cell union in mating pairs in the ciliate Euplotes raikovi is controlled by a system of multiple mating types which are inherited with alleles codominant at the genetic locus mat and expressed via diffusible mating pheromones. The mating pheromones Er-2, Er-3, and Er-11 were purified from cells homozygous for the mat-2, mat-3, and mat-11 alleles, respectively. These pheromones are proteins of similar Mr (11,000-12,000) and acidity (pI 3.7-4.0) and are active at a concentration that varies from 2.9 X 10(-12) to 1.2 X 10(-11) M. Data on amino acid composition revealed that an unusually high amount of cysteine (12-15.7%) and poor contents of basic amino acids are common to every pheromone. On the basis of this uniformity in the main biochemical traits, which also holds for the previously purified pheromone Er-1, it was concluded that E. raikovi mating pheromones are members of a family of proteins structurally diversified from each other to varying extents.

Alleles↗

PubMed1986

Purification to apparent homogeneity of the mating pheromone of mat-1 homozygous Euplotes raikovi.

Mating type-specific mating pheromones autonomously released into the environment control cell-cell union in conjugation of the marine ciliate Euplotes raikovi. The mating pheromone, termed euplomone r-1, of cells of the homozygous mating type determined by the mat-1/mat-1 genotype was purified by means of a three-stage purification procedure which provides a yield of 79%. Starting from 10 liters of supernatant, 3.3 mg of euplomone r-1 were regularly recovered. Euplomone r-1 was identified with a protein showing a molecular weight of 12,000, an isoelectric point of 3.7, and unusually high contents of cysteine (15%) and tyrosine (5.8%). It appeared active at a concentration of 3.4 +/- 0.5 X 10(-12) M. Carbohydrate and a small colored compound, not yet identified, occurred in association with partially purified euplomone r-1 samples, whereas they were not detected in samples of the final product of purification. An acidic shift was apparently capable of destroying the carbohydrate/euplomone r-1 association, suggesting that it does not involve covalent bonds.

Amino Acids↗

PubMed1986

The ciliate Euplotes raikovi heterozygous at the mat genetic locus coreleases two individual species of mating pheromone: Genetic and biochemical evidence.

Euplotes raikovi of the wild-type strain 13 was found to be heterozygous (mat-1/mat-2) for the genetic locus mat, which is supposed to control the mating-type specificity of freely released mating pheromones ("euplomones"), and capable of yielding the two types of corresponding homozygotes (mat-1/mat-1, mat-2/mat-2). The results of euplomone purification, performed in parallel on Euplotes of the three different genotypes, showed that the heterozygous cells corelease two mat-specific euplomones (namely, euplomone r 1 associated with mat-1 and euplomone r 2 associated with mat-2), while the homozygous cells release either one of the two types according to their genotype. The two euplomones coreleased by the mat heterozygous cells were resolved as separate molecular species present in different relative amounts (euplomone r 1, approximately 70%; euplomone r 2, approximately 30%). A closely similar degree of eccentricity in the euplomone r 1/euplomone r 2 production was again found between the two homozygous cell types. It was concluded that the alleles mat-1 and mat-2 exhibit a relationship of nondominance: the heterozygote apparently behaves as the simple combination of the two corresponding homozygotes. It was inferred that the observed quantitative variations in the production of the different euplomones may be the result of a differential mat gene-type amplification occurring during the development of the cell somatic macronucleus.

Journal Article↗

PubMedDate not supplied

Specific and common epitopes in mating pheromones of Euplotes raikovi revealed by monoclonal antibodies.

Polypeptide mating pheromones Er-1 and Er-2, purified from the supernatant of Euplotes raikovi cultures of mating type I and mating type II, respectively, were used to immunize mice and obtain monoclonal antibodies. Five hybridoma clones producing antibodies specific to the mating pheromones were selected. They were analyzed for immunospecificity by immunoperoxidase assay, immunoblotting, and for their efficacy in inhibition of mating pheromone activity. Monoclonal antibodies from two hybridoma clones recognized only the mating pheromone used as antigen: those from the other three clones reacted, to comparable extents, with both mating pheromones. On the basis of these results it was assumed that two immunogenic sites exist in Er-1 and Er-2, one specific and the other common to both mating pheromones.

Animals↗

PubMedDate not supplied

Self from nonself discrimination in single-celled eukaryotes.

A mechanism for self recognition has been proposed to control the mating type interactions in the marine ciliate Euplotes raikovi. The relevant molecules involved in this mechanism are the mating pheromones inherited via high-multiple alleles codominant at the Mendelian locus mat. Four of these mating pheromones have been isolated, purified, and characterized. They are relatively small and acidic proteins (Mr, 11,000-12,000; pI, 3.7-4.0), which are still active at the concentration of approximately 10(-12) molar. Data, yet preliminary, of amino acid sequencing of the purified mating pheromones showed extensive structural homologies and the common presence of the aspartic acid at the amino terminal.

Animals↗