Topography of cell wall lytic enzyme in Chlamydomonas reinhardtii: form and location of the stored enzyme in vegetative cell and gamete.
Chlamydomonas lytic enzyme of the cell wall (gamete wall-autolysin) is responsible for shedding of cell walls during mating of opposite mating-type gametes. This paper reports some topographic aspects of lytic enzyme in cells. Both vegetative and gametic cells contain the same wall lytic enzyme. The purified enzyme is a glycoprotein with an apparent molecular mass of 67 kD by gel filtration and 62 kD by SDS PAGE, and is sensitive to metal ion chelators and SH-blocking agents. These properties are the same as those of the gamete wall-autolysin released into the medium by mating gametes. However, the storage form of the enzyme proves to be quite different between the two cell types. In vegetative cells, the lytic enzyme is found in an insoluble form in cell homogenates and activity is released into the soluble fraction only by sonicating the homogenates or freeze-thawing the cells, whereas gametes always yield lytic activity in the soluble fractions of cell homogenates. When vegetative cells are starved for nitrogen, the storage form of enzyme shifts from its vegetative state to gametic state in parallel with the acquisition of mating ability. Adding nitrogen to gametes converts it to the vegetative state concurrently with the loss of mating ability. We also show that protoplasts obtained by treatment of vegetative cells or gametes with exogenously added enzyme have little activity of enzyme in the cell homogenates, suggesting that lytic enzyme is stored outside the plasmalemma. When the de-walled gametes or gametes of the wall-deficient mutant, cw-15, of opposite mating types are mixed together, they mate normally but the release of lytic enzyme into the medium is practically negligible. When the de-walled vegetative cells are incubated, the lytic enzyme is again accumulated in the cells after the wall regeneration is almost complete.
D000135:Acid Phosphatase / Q000302:isolation & purification / Q000378:metabolism; D000581:Amidohydrolases / Q000378:metabolism*; D001329:Autolysis; D002696:Chlamydomonas / Q000201:enzymology* / Q000254:growth & development; D007700:Kinetics; D008970:Molecular Weight; D009154:Mutation; D009238:N-Acetylmuramoyl-L-alanine Amidase / Q000302:isolation & purification / Q000378:metabolism*; D011523:Protoplasts / Q000201:enzymology; D013045:Species Specificity
D000135:Acid Phosphatase; D000581:Amidohydrolases; D009238:N-Acetylmuramoyl-L-alanine Amidase