L-Amino-acid oxidase from the Malayan pit viper Calloselasma rhodostomaComparative sequence analysis and charaterization of active and inactive forms of the enzyme

Macheroux, Peter; Seth, Oliver; Bollschweiler, Claus; Schwarz, Margarete; Kurfürst, Manfred; Au, Lo‐Chun; Ghisla, Sandro

doi:10.1046/j.1432-1033.2001.02042.x

Cited by 39 publications

(51 citation statements)

References 14 publications

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“…Thus, like all LAAO enzymes described thus far, IL4I1 is secreted. 9,17 Second, IL4I1 preferentially hydrolyzed L-phenylalanine and its activity was not restricted to acidic pH, being also active at a pH approaching neutrality. Surprisingly, some of these results differed from data published by Mason et al for the mouse enzyme, although we used the same experimental conditions to test the enzymatic activity.…”

Section: Discussionmentioning

confidence: 99%

Human IL4I1 is a secreted l-phenylalanine oxidase expressed by mature dendritic cells that inhibits T-lymphocyte proliferation

Boulland

Marquet

Molinier‐Frenkel

et al. 2007

Blood

159

177

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Interleukin-4-induced gene 1 (IL4I1) was first described as a B-cell IL4-inducible gene and is highly expressed in primary mediastinal B-cell lymphomas. We established stable HEK293 clones expressing human and mouse IL4I1 to examine their biochemical properties and function. Both proteins were secreted into the culture medium, and we observed the secretion of endogenous human IL4I1 (hIL4I1) protein in a mediastinal lymphoma B-cell line, MedB-1. We showed that IL4I1 has L-amino acid oxidase activity, optimal at physiological pH and primarily directed toward phenylalanine. Immunohistochemical analysis of secondary lymphoid organs showed staining of germinal center macrophages and inflammatory myeloid cells. In vitro, functional enzyme was highest in mature dendritic cells (DCs), suggesting a role in antigenpresenting cell/T-lymphocyte cross-talk. Indeed, hIL4I1 inhibited the proliferation of CD3-stimulated T lymphocytes with a similar effect on CD4 ؉ and CD8 ؉ T cells. IntroductionInterleukin-4 (IL4) is the master cytokine for T-helper type 2 (Th2) lymphocyte differentiation and function. Along with its important role in B-lymphocyte stimulation by T-helper cells in germinal centers of secondary lymphoid organs, it also regulates the growth and function of many different cell types of the immune system. 1 IL4 transcriptional effects are primarily mediated by the signalization and transcription factor 6 (STAT6), which is responsible for the induction and repression of multiple target genes. 2 An immediate-early IL4-inducible gene called interleukin fourinduced gene 1 (FIG1/IL4I1) has first been described in the mouse 3 and subsequently characterized in human B cells. 4 IL4I1 mRNA expression is restricted to lymphoid tissues, with the highest levels found in lymph nodes and spleen. 4,5 IL4-activated murine B cells express IL4I1 within 2 hours of stimulation. We have demonstrated that high levels of expression of this gene are characteristic of primary mediastinal large B-cell lymphoma (PMBL), a specific subtype of diffuse large B-cell lymphoma. 5 The high expression of IL4I1 by PMBL may result from the constitutive activation of STAT6 in these tumors. 6 Both human and mouse IL4I1 mRNA sequences encode a protein containing a putative signal peptide indicative of potential secretion and a large central domain, highly homologous to flavin-containing amino acid oxidases. 3,4 While sharing high similarity over the majority of the sequence (547 amino acids of the 567 in the human sequence), the human and mouse proteins diverge substantially at their C-terminal region. 4 The IL4I1 protein shares the highest similarity with proteins presenting L-amino acid oxidase (LAAO) activity, such as snake venom LAAO,7,8 and mouse milk LAAO. 9 Recently, Mason et al have demonstrated that mouse IL4I1 (mIL4I1) possesses an LAAO activity toward aromatic amino acids. The enzyme was mostly active at acidic pH, in contrast to other known members of the LAAO family, and its expression was restricted primarily to lysosomes. 10 In this work, w...

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Section: Discussionmentioning

confidence: 99%

Human IL4I1 is a secreted l-phenylalanine oxidase expressed by mature dendritic cells that inhibits T-lymphocyte proliferation

Boulland

Marquet

Molinier‐Frenkel

et al. 2007

Blood

159

177

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“…L-Amino acid oxidase (LAAO) from the venom of the Malayan pit viper C. rhodostoma was obtained as described (Macheroux et al, 2001).…”

Section: General Chemicals Reagents and Enzymesmentioning

confidence: 99%

Induction of apoptosis in yeast by L‐amino acid oxidase from the Malayan pit viper Calloselasma rhodostoma

Ande

Fussi

Knauer

et al. 2008

Yeast

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Here we report for the first time that L-amino acid oxidase (LAAO), a major component of snake venom, induces apoptosis in yeast. The causative agent for induction of apoptosis has been shown to be hydrogen peroxide, produced by the enzymatic activity of LAAO. However, the addition of catalase, a specific hydrogen peroxide scavenger, does not prevent cell demise completely. Intriguingly, depletion of leucine from the medium by LAAO and the interaction of LAAO with yeast cells are shown to be the major factors responsible for cell demise in the presence of catalase.

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“…Glucose oxidase and catalase were from Boehringer Mannheim. L-Amino acid oxidase was obtained as described elsewhere (26). Ferricenium hexafluorophosphate (FcPF6, Aldrich) was prepared according to Lehman et al (27).…”

Section: Methodsmentioning

confidence: 99%

Solvent Isotope Effects in Reactions of Human Medium-Chain Acyl-CoA Dehydrogenase Active Site Mutants

2007

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Glu376, the base involved in substrate RH + abstraction at the active center of medium-chain acyl-CoA dehydrogenase (MCAD), has been mutated to Gln and Gly. The mutants are active; however, their rates of dehydrogenation are lowered by approximately 5 orders of magnitude. Binding of the substrate octanoyl-CoA to Glu376Gln-MCAD involves (at least) two steps. The ensuing dehydrogenation reaction that corresponds to reduction of the flavin cofactor also occurs in two phases. These are interpreted to consist of a first, reversible step, followed by a slower, practically irreversible one. For Glu376Gln-MCAD, the log of the rates of dehydrogenation increases linearly with pH (slope ) 1) in the pH range of 6-10, suggesting HO -as a reactant. The rates of the same reactions in D 2 O have the same pD profile and reflect a solvent kinetic isotope effect (SKIE) of ≈8.5. Glu376Gln+Glu99Gly-MCAD (studied to assess the role of Glu99 also present at the bottom of the active center cavity) has activities and activity profiles similar to those of Glu376Gln-MCAD. This excludes Glu99 as the active center base for Glu376Gln-MCAD catalysis. Proton inventories for the two phases of the dehydrogenation reaction were investigated at 4 and 25°C. The inventories at 25°C reflect a SKIE of ≈4.5; the profiles are "bowlshaped", in which a transition-state contribution predominates. The profiles for the 4°C reaction are very unusual. That for the first phase can be analyzed on a two-step model with one step (80% rate-limiting) having a conformational reorganization with an isotope effect of 90-100, from small isotope effects at many protein sites, and the other step (20% rate-limiting) having an inverse isotope effect of ca. 2, characteristic of the reaction of hydroxide ion as a base. For the second phase, only a contribution from many protein sites with a KIE of ≈4.5 is observed. The results are compatible with a very rigid active site framework that must undergo rearrangements for dehydrogenation to take place, and specifically to allow access of HO -, the reactant that must neutralize the H + abstracted from the RC-H substrate. The large isotope effects are attributed to the changes in state of several H-bonds that occur during the process.

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L-Amino-acid oxidase from the Malayan pit viper Calloselasma rhodostomaComparative sequence analysis and charaterization of active and inactive forms of the enzyme

Cited by 39 publications

References 14 publications

Human IL4I1 is a secreted l-phenylalanine oxidase expressed by mature dendritic cells that inhibits T-lymphocyte proliferation

Human IL4I1 is a secreted l-phenylalanine oxidase expressed by mature dendritic cells that inhibits T-lymphocyte proliferation

Induction of apoptosis in yeast by L‐amino acid oxidase from the Malayan pit viper Calloselasma rhodostoma

Solvent Isotope Effects in Reactions of Human Medium-Chain Acyl-CoA Dehydrogenase Active Site Mutants

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