Intracellular distribution of nitrogen during synchronous growth of Chlorella pyrenoidosa

A continuous dilution method for the mass culture of microorganisms was developed which yielded 3x as much cellular material as previous culturing methods for C. pyrenoidosa in the same time period and in less than one-tenth the culture volume. Cellular growth parameters increased exponentially throughout the cell cycle of C. pyrenoidosa when the new mass culturing technique was utilized.

The levels of nitrogen in different cellular fractions were estimated during the cell cycle (i.e. lipid-plus chlorophyll-N; TCA soluble-N; individual free-, peptide-, and protein-amino acid residues; nucleotide-N; DNA-N; RNA-N).

The amino acid distribution within the cellular protein remained nearly constant during cellular development. Protein-N accounted for approximately 60% of the total cellular-N throughout the cell cycle.

The free amino acids exhibited a variety of trends in change of level during the cell cycle. Free-alanine, -lysine, -serine, -glycine, -arginine, and -glutamate were present at relatively high levels. Increase in level of peptide-N during early cellular development resulted largely from increase in levels of peptide-arginine, -glutamate, and -lysine. The other peptide amino acids exhibited a variety of different trends.

RNA-N demonstrated exponential accumulation throughout cellular development with a reduced rate of accumulation during cellular division. DNA-N increased during most of the cell cycle at a lower exponential rate; however, the rate of DNA-N accumulation increased abruptly at the onset of nuclear division.

Reduced glutathione (GSH) was found to be the predominant acid-soluble sulfhydryl-containing compound throughout the cell cycle. Its trend (as % of total cellular-N) was similar to that of TCA-soluble sulfhydryl-containing compounds in synchronized sea urchin eggs.

GSH exhibited properties similar to certain compounds tentatively identified as sulfur-containing nucleotide-peptides by previous workers. Norit-A adsorption studies indicated that nucleotide-peptides were not present in the TCA extracts of C, pyrenoidosa.

During the period 0.4-0.9 fraction of the cell cycle GSH was synthesized at the expense of cellular protein. A hypothetical scheme was presented to account for both the origin of GSH prior to nuclear division and the proposed regulation of mitotic apparatus formation through acid-soluble sulfhydryl compounds.