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Updated: Mar 28, 2026
Quantitative Methods to Study Protein Arginine Methyltransferase 1-9 Activity in Cells
Published on: August 7, 2021
Biochemical and biological activity of arginine deiminase from Streptococcus pyogenes M22
Eleonora A Starikova1, Alexey V Sokolov1,2, Anna Yu Vlasenko1
1a Institute of Experimental Medicine, 12 Pavlov Street, St. Petersburg, 197376, Russia.
Abstract:
Streptococcus pyogenes (group A Streptococcus; GAS) is an important gram-positive extracellular bacterial pathogen responsible for a number of suppurative infections. This micro-organism has developed complex virulence mechanisms to avoid the host's defenses. We have previously reported that SDSC from GAS type M22 causes endothelial-cell dysfunction, and inhibits cell adhesion, migration, metabolism, and proliferation in a dose-dependent manner, without affecting cell viability. This work aimed to isolate and characterize a component from GAS type M22 supernatant that suppresses the proliferation of endothelial cells (EA.hy926). In the process of isolating a protein possessing antiproliferative activity we identified arginine deiminase (AD). Further study showed that this enzyme is most active at pH 6.8. Calculating Km and Vmax gave the values of 0.67 mmol·L(-1) and 42 s(-1), respectively. A distinctive feature of AD purified from GAS type M22 is that its optimum activity and the maximal rate of the catalytic process is close to neutral pH by comparison with enzymes from other micro-organisms. AD from GAS type M22 suppressed the proliferative activity of endothelial cells in a dose-dependent mode. At the same time, in the presence of AD, the proportion of cells in G0/G1 phase increased. When l-Arg was added at increasing concentrations to the culture medium containing AD (3 μg·mL(-1)), the enzyme's capacity to inhibit cell proliferation became partially depressed. The proportion of cells in phases S/G2 increased concomitantly, although the cells did not fully recover their proliferation activity. This suggests that AD from GAS type M22 has potential for the suppression of excessive cell proliferation.
Insights
Streptococcus pyogenes releases arginine deiminase (AD), an enzyme that suppresses endothelial cell proliferation. This bacterial enzyme shows potential for controlling excessive cell growth by affecting cell cycle progression.
Area of Science:
- Microbiology
- Biochemistry
- Cell Biology
Background:
- Streptococcus pyogenes (group A Streptococcus; GAS) is a gram-positive bacterium causing suppurative infections.
- GAS employs complex virulence mechanisms to evade host defenses, including factors that affect host cell functions.
- Previous studies indicated that GAS type M22 supernatant inhibits endothelial cell functions like proliferation.
Purpose of the Study:
- To isolate and characterize a component from GAS type M22 supernatant with endothelial cell antiproliferative activity.
- To investigate the properties and mechanism of action of the identified antiproliferative enzyme.
Main Methods:
- Isolation and purification of a protein with antiproliferative activity from GAS type M22 supernatant.
- Enzyme kinetics analysis of arginine deiminase (AD), including determination of Km and Vmax.
- Assessment of AD's effect on endothelial cell proliferation and cell cycle distribution (G0/G1, S/G2 phases).
- Evaluation of the effect of L-arginine supplementation on AD-mediated inhibition of cell proliferation.
Main Results:
- Arginine deiminase (AD) was identified as the component suppressing endothelial cell proliferation.
- GAS M22 AD exhibits optimal activity near neutral pH (pH 6.8), with Km of 0.67 mmol·L(-1) and Vmax of 42 s(-1).
- AD inhibited endothelial cell proliferation in a dose-dependent manner, increasing the proportion of cells in G0/G1 phase.
- L-arginine partially reversed AD's inhibitory effect on proliferation, suggesting arginine depletion as a mechanism.
Conclusions:
- Arginine deiminase from GAS type M22 is a key enzyme that suppresses endothelial cell proliferation.
- The enzyme's activity is influenced by pH and substrate availability (L-arginine).
- GAS M22 AD demonstrates potential for therapeutic applications in controlling excessive cell proliferation.
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