Structural basis of enzyme activity regulation by the propeptide of l-lysine α-oxidase precursor from Trichoderma viride

Kawamura, Masaki; Ito, Norihisa; Matsumoto, Yuya; Saito, Masaya; Tamura, Takashi; Kusakabe, Hitoshi; Inagaki, Kenji; Imada, Katsumi

doi:10.1016/j.yjsbx.2021.100044

Cited by 4 publications

(4 citation statements)

References 24 publications

(18 reference statements)

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“…Biochemical, structural, and computational analysis of AncLLysO2 indicated that the enzyme is a less promiscuous l -Lys α-oxidase having no pro-sequence; LAAO activity is often toxic for living organisms, so to reduce the activity under in vivo conditions, several of the LAAOs hinder the substrate entrance pathway by the pro-sequence. , The properties were different from other LLysOs; catalytic promiscuity (oxidase and mono-oxygenase activity) was confirmed in AncLLysO and L-LOX/MOG, and l -Lys α-oxidase from Trichoderma viride (TvLysO) matured by cleaving the pro-sequence . Sequence identity among the enzymes was <32% (Table S4), suggesting that the enzymes have unique substrate recognition mechanisms.…”

Section: Resultsmentioning

confidence: 99%

“… 44 , 45 The properties were different from other LLysOs; catalytic promiscuity (oxidase and mono-oxygenase activity) was confirmed in AncLLysO and L-LOX/MOG, and l -Lys α-oxidase from Trichoderma viride (TvLysO) matured by cleaving the pro-sequence. 46 Sequence identity among the enzymes was <32% ( Table S4 ), suggesting that the enzymes have unique substrate recognition mechanisms.…”

Section: Resultsmentioning

confidence: 99%

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Reaction Mechanism of Ancestral l-Lys α-Oxidase from Caulobacter Species Studied by Biochemical, Structural, and Computational Analysis

et al. 2022

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The flavin-dependent amine oxidase superfamily contains various l-amino acid oxidases (LAAOs) bearing different substrate specificities and enzymatic properties. LAAOs catalyze the oxidation of the α-amino group of l-amino acids (L-AAs) to produce imino acids and H2O2. In this study, an ancestral l-Lys α-oxidase (AncLLysO2) was designed utilizing genome-mined sequences from the Caulobacter species. The AncLLysO2 exhibited high specificity toward l-Lys; the k cat/K m values toward l-Lys were one and two orders larger than those of l-Arg and l-ornithine, respectively. Liquid chromatography–high resolution mass spectrometry analysis indicated that AncLLysO2 released imino acid immediately from the active site after completion of oxidation of the α-amino group. Crystal structures of the ligand-free, l-Lys- and l-Arg-bound forms of AncLLysO2 were determined at 1.4–1.6 Å resolution, indicating that the active site of AncLLysO2 kept an open state during the reaction and more likely to release products. The structures also indicated the substrate recognition mechanism of AncLLysO2; ε-amino, α-amino, and carboxyl groups of l-Lys formed interactions with Q357, A551, and R77, respectively. Biochemical and molecular dynamics simulation analysis of AncLLysO2 indicated that active site residues that indirectly interact with the substrate are also important to exhibit high activity; for example, the aromatic group of Y219 is important to ensure that the l-Lys substrate is placed in the correct position to allow the reaction to proceed efficiently. Taken together, we propose the reaction mechanism of AncLLysO2.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Reaction Mechanism of Ancestral l-Lys α-Oxidase from Caulobacter Species Studied by Biochemical, Structural, and Computational Analysis

et al. 2022

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“…The l -lysine α-oxidase (LO) from Trichoderma harzianum is a flavoprotein from the family of l -amino acid oxidases (LAAOs) [ 32 ], an enzyme family extensively studied in medicine for their cytotoxic effects in different tumor cells. However, the large-scale expression and isolation in prokaryotic hosts such as E. coli [ 17 ] is a challenge for the development of commercial recombinant LAAOs, mainly due to the formation of inclusion bodies [ 33 ], production of the inactive [ 34 ], and/or low yield [ 35 ]. Another issue with the heterologous expression of these enzymes is that they are usually expressed as propeptide precursors to protect the host organism from the LAAO toxicity [ 35 ].…”

Section: Discussionmentioning

confidence: 99%

The recombinant l-lysine α-oxidase from the fungus Trichoderma harzianum promotes apoptosis and necrosis of leukemia CD34 + hematopoietic cells

Costa,

Silva,

Guimarães

et al. 2024

Microb Cell Fact

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Background In hematologic cancers, including leukemia, cells depend on amino acids for rapid growth. Anti-metabolites that prevent their synthesis or promote their degradation are considered potential cancer treatment agents. Amino acid deprivation triggers proliferation inhibition, autophagy, and programmed cell death. l-lysine, an essential amino acid, is required for tumor growth and has been investigated for its potential as a target for cancer treatment. l-lysine α-oxidase, a flavoenzyme that degrades l-lysine, has been studied for its ability to induce apoptosis and prevent cancer cell proliferation. In this study, we describe the use of l-lysine α-oxidase (LO) from the filamentous fungus Trichoderma harzianum for cancer treatment. Results The study identified and characterized a novel LO from T. harzianum and demonstrated that the recombinant protein (rLO) has potent and selective cytotoxic effects on leukemic cells by triggering the apoptotic cascade through mitochondrial dysfunction. Conclusions The results support future translational studies using the recombinant LO as a potential drug for the treatment of leukemia.

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“…The L-lysine α-oxidase (LO) from Trichoderma harzianum is a avoprotein from the family of L-amino acid oxidases (LAAOs) [32], an enzyme family extensively studied in medicine for their cytotoxic effects in different tumor cells. However, a challenge for the development of commercial recombinant LAAOs is the large-scale expression and isolation in prokaryotic hosts such as E. coli [33], mainly due to the formation of inclusion bodies [34], production of the inactive [35], and/or low yield [51]. Another di culty in the heterologous expression is that they are usually expressed as propeptide precursors, rendering the host organism protection from the LAAO toxicity [36].…”

Section: Discussionmentioning

confidence: 99%

The recombinant L-Lysine α-oxidase from the fungus Trichoderma harzianum promotes apoptosis and necrosis of leukemia CD34+ hematopoietic cells

Costa

Silva

Guimarães

et al. 2023

Preprint

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Background In hematologic cancers, including leukemia, cells depend on amino acids for rapid growth. Anti-metabolites that prevent their synthesis or promote their degradation are considered potential cancer treatment agents. Amino acid deprivation triggers proliferation inhibition, autophagy, and programmed cell death. Lysine, an essential amino acid, is required for tumor growth and has been investigated for its potential as a target for cancer treatment. L-lysine-α-oxidase, a flavoenzyme that degrades lysine, has been studied for its ability to induce apoptosis and prevent cancer cell proliferation. In this study, we describe the use of L-lysine oxidase (LO) from the filamentous fungus Trichoderma harzianum for cancer treatment. Results The study identified and characterized a novel LO from T. harzianum and demonstrated that the recombinant protein (rLO) has potent and selective cytotoxic effects on leukemic cells by triggering the apoptotic cascade through mitochondrial dysfunction. Conclusions The results support future translational studies using the recombinant LO as a potential drug for the treatment of leukemia.

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Structural basis of enzyme activity regulation by the propeptide of l-lysine α-oxidase precursor from Trichoderma viride

Abstract: Graphical abstract

Cited by 4 publications

References 24 publications

Reaction Mechanism of Ancestral l-Lys α-Oxidase from Caulobacter Species Studied by Biochemical, Structural, and Computational Analysis

Reaction Mechanism of Ancestral l-Lys α-Oxidase from Caulobacter Species Studied by Biochemical, Structural, and Computational Analysis

The recombinant l-lysine α-oxidase from the fungus Trichoderma harzianum promotes apoptosis and necrosis of leukemia CD34 + hematopoietic cells

The recombinant L-Lysine α-oxidase from the fungus Trichoderma harzianum promotes apoptosis and necrosis of leukemia CD34+ hematopoietic cells

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