S-Adenosylmethionine synthetase from human lymphocytes. Purification and characterization

Area of Science:

• Biochemistry
• Enzymology

Background:

• S-Adenosylmethionine synthetase (MAT) is crucial for cellular methylation.
• Understanding MAT's properties is vital for comprehending leukemia cell metabolism.

Purpose of the Study:

• To purify and characterize S-Adenosylmethionine synthetase from human chronic lymphocytic leukemia (CLL) cells.
• To elucidate the enzyme's subunit composition, kinetic mechanism, and cofactor requirements.

Main Methods:

• Enzyme purification to homogeneity.
• Equilibrium sedimentation and SDS-PAGE for molecular weight and subunit analysis.
• Kinetic studies including initial velocity and product inhibition assays.

Main Results:

• Purified enzyme with a native molecular weight of 185,000 and complex subunit composition (alpha, alpha', beta).
• Demonstrated ordered Bi Ter mechanism for substrate addition (ATP, L-methionine) and random release of pyrophosphate/orthophosphate.
• Identified Mg2+ as an essential activator and characterized kinetic parameters (Km, Kia).
• Observed tripolyphosphatase activity stimulated by S-adenosylmethionine.

Conclusions:

• The purified S-Adenosylmethionine synthetase from CLL cells possesses a distinct subunit structure and complex kinetic mechanism.
• Kinetic and cofactor data provide insights into the enzyme's function in leukemia cells.
• The enzyme's characteristics may offer potential therapeutic targets in chronic lymphocytic leukemia.