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<p>Intracellular L-methionine γ-lyase (MGL) from <i>Klebsiella oxytoca</i> BLM-1 was produced and optimized using a combination of One-Factor-at-a-Time and Response Surface Methodology. Optimal culture conditions, such as pH 9.0, 3% (w/v) lactose, and 1.02% (w/v) malt extract, resulted in the highest intracellular MGL activity (0.235 U/mL), representing a 1.13-fold improvement over initial conditions, with a total yield of 40.80 U from a 2 L optimized broth. Purification using Octyl-Sepharose chromatography produced a highly active multimeric enzyme (~ 250&#xa0;kDa) with 0.384 U/mL activity, which was confirmed as a heteromeric complex (~ 63&#xa0;kDa and ~ 117&#xa0;kDa subunits) by SDS-PAGE. The enzyme displayed strong cytotoxic activity toward methionine-dependent cancer cell lines, with IC₅₀ values of 0.023 U for HepG2 and 0.0045 U for A549, while exerting minimal effects on HEK-293 cells. Molecular docking revealed that L-methionine binds to the MGL active site with a binding energy of − 6.5&#xa0;kcal/mol, and protein–protein docking identified favorable interactions with key methionine pathway enzymes, including METAP2 (− 902.3), MAT2A (− 895.1), and SAHH (− 890.9). These findings highlight the successful optimization of MGL production, its effective purification, and its significant anticancer potential, providing a strong foundation for its development as a therapeutic enzyme for methionine-dependent tumors.</p>

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Integrated production and protein–protein docking analysis of L-methionase from Klebsiella oxytoca for enzyme-based anticancer therapy via methionine starvation and epigenetic modulation

  • 期刊论文

  • 发表日期：

    2025年9月9日

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  • Bhupender Sharma,
  • Vivek Chauhan,
  • Vivek Kumar Dhiman,
  • Rakesh Kumar,
  • Gaytri Mahajan,
  • Sukhdev Singh,
  • Kanta Sashi,
  • Anjali Kashwal,
  • Shamsher S. Kanwar

摘要

Intracellular L-methionine γ-lyase (MGL) from Klebsiella oxytoca BLM-1 was produced and optimized using a combination of One-Factor-at-a-Time and Response Surface Methodology. Optimal culture conditions, such as pH 9.0, 3% (w/v) lactose, and 1.02% (w/v) malt extract, resulted in the highest intracellular MGL activity (0.235 U/mL), representing a 1.13-fold improvement over initial conditions, with a total yield of 40.80 U from a 2 L optimized broth. Purification using Octyl-Sepharose chromatography produced a highly active multimeric enzyme (~ 250 kDa) with 0.384 U/mL activity, which was confirmed as a heteromeric complex (~ 63 kDa and ~ 117 kDa subunits) by SDS-PAGE. The enzyme displayed strong cytotoxic activity toward methionine-dependent cancer cell lines, with IC₅₀ values of 0.023 U for HepG2 and 0.0045 U for A549, while exerting minimal effects on HEK-293 cells. Molecular docking revealed that L-methionine binds to the MGL active site with a binding energy of − 6.5 kcal/mol, and protein–protein docking identified favorable interactions with key methionine pathway enzymes, including METAP2 (− 902.3), MAT2A (− 895.1), and SAHH (− 890.9). These findings highlight the successful optimization of MGL production, its effective purification, and its significant anticancer potential, providing a strong foundation for its development as a therapeutic enzyme for methionine-dependent tumors.