| Research Abstract |
This project was started to make clear the mutual correlation among the heterogeneous hemoglobin components and the structure of the hemoglobin genes in Paramecium.
We found that, using 6 stocks in 3 different species of Paramecium, 1) relative amounts of hemoglobin components shows a definite change during the cell growth and by the temperature of the culture, 2) the changes occur between the major hemoglobin component and submajor one, 3) the former in a cell shows faster migration by PAGE, smaller in molecular weight and lower in pI than the latter, and 4) both of the components increase or decrease inversely their amounts each other.
It was confirmed that the major hemoglobin components (Hb10) from 15 stocks in five sibling species of P. caudatum are dividable into three types, as A, B, and A + B, although their pI values fall into a narrow range measuring about pH 3.9. The submajor hemoglobin component in this species is also divided into four variants, showing the pI value of pH 9.8-10.5.
We determined the nucleotide sequence of the cDNA encoding the A-type of Hb10. The results obtained indicate that the sequence contains four TAA codons, which are known to act as a termination signal in the most of eukaryotes but encode Gln for ciliates, and that the Hb has an unusual structure which is composed of 116 amino acid residues. Analysis of the genomic DNA from macronuclei made clear that the gene is organized with two exons interrupted by one small intron, which is composed of only 23 bp at the position corresponding to the second intron found in plant Hb genes. We also clarified that the gene encoding the B-type of Hb10 are identical to that of the A-type Hb10, in an exception of the replacements of five out of 116 amino acid residues.
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