Intracellular distribution of nitrogen during synchronous growth of Chlorella pyrenoidosa

Abstract

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

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.