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Detecting Cortex Fragments During Bacterial Spore Germination
Published on: June 25, 2016
Acid protease activity during germination of microcysts of the cellular slime mold Polysphondylium pallidum
Abstract:
Extracts of dormant microcysts of Polysphondylium pallidum demonstrate pH optima for the hydrolysis of casein at 3.5 and 6.0. During germination the intracellular pH 6.0 caseinolytic specific activity does not change significantly. The pH 6.0 protease is also active on azo-albumin, revealing the same developmental pattern with this substrate. Both acid protease activities are excreted during the germination process. Addition of purified nonspecific protease to cultures speeds up germination, suggesting that the excreted protease may play a role in removal of the microcyst wall. When cycloheximide is added to cultures, complete germination (emergence) is stopped whereas the pH 6.0 protease activity still accumulates to between 50 and 60% of the maximum control activity. Although this suggests that post-translational controls might mediate the accumulation of a portion of the pH 6.0 protease increase, mixing and dilution experiments with cell extracts do not reveal the differential presence of soluble activators or inhibitors of this activity at different developmental stages. The presence of tightly bound enzyme-inhibitor complexes for protease B in dormant microcysts has not been ruled out and is currently under study.
Insights
Polysphondylium pallidum microcysts release proteases during germination, aiding in microcyst wall breakdown. Cycloheximide inhibits germination but not protease accumulation, suggesting post-translational regulation.
Area of Science:
- Cell Biology
- Biochemistry
- Developmental Biology
Background:
- Polysphondylium pallidum is a social amoeba.
- Microcysts are dormant structures resistant to environmental stress.
- Germination involves the emergence of vegetative cells from microcysts.
Purpose of the Study:
- To investigate the role of proteases in the germination of Polysphondylium pallidum microcysts.
- To characterize the pH optima and activity of proteases during germination.
- To explore the regulation of protease activity during microcyst germination.
Main Methods:
- Enzyme activity assays on casein and azo-albumin.
- pH optimum determination for protease hydrolysis.
- Effect of cycloheximide on germination and protease activity.
- Mixing and dilution experiments with cell extracts.
Main Results:
- Two pH optima for casein hydrolysis were identified at pH 3.5 and 6.0.
- Protease activity is excreted during germination, with the pH 6.0 protease showing similar developmental patterns on azo-albumin.
- Exogenous protease accelerates germination, suggesting a role in microcyst wall degradation.
- Cycloheximide halts complete germination but allows pH 6.0 protease accumulation, indicating post-translational control.
Conclusions:
- Excreted proteases likely facilitate microcyst wall removal during Polysphondylium pallidum germination.
- Post-translational mechanisms may regulate the accumulation of pH 6.0 protease.
- Further investigation is needed to rule out enzyme-inhibitor complexes in dormant microcysts.
