Metal coordination by L-amino acid oxidase derived from flounder Platichthys stellatus is structurally essential and regulates antibacterial activity

Kasai, Kosuke; Ito, Yudai; Nitta, Akihide; Ariyoshi, Kentaro; Nakamura, Toshiya; Miura, Tomisato

doi:10.1007/s00253-020-10914-3

Cited by 5 publications

(3 citation statements)

References 33 publications

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“…BT’s 3D structure was created with SWISS-MODEL with a global model quality estimation (GMQE) score of 0.83. A GMQE score greater than 0.70 is regarded as a trustworthy predictor in general [ 26 ]. In addition, the crystal structure of BT was shown to have a similarity identity of 80% with template protein.…”

Section: Resultsmentioning

confidence: 99%

Homology Modeling, Molecular Docking, Molecular Dynamic Simulation, and Drug-Likeness of the Modified Alpha-Mangostin against the β-Tubulin Protein of Acanthamoeba Keratitis

et al. 2022

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Acanthamoeba species are capable of causing amoebic keratitis (AK). As a monotherapy, alpha-mangostin is effective for the treatment of AK; however, its bioavailability is quite poor. Moreover, the efficacy of therapy is contingent on the parasite and virulent strains. To improve readiness against AK, it is necessary to find other derivatives with accurate target identification. Beta-tubulin (BT) has been used as a target for anti-Acanthamoeba (A. keratitis). In this work, therefore, a model of the BT protein of A. keratitis was constructed by homology modeling utilizing the amino acid sequence from NCBI (GenBank: JQ417907.1). Ramachandran Plot was responsible for validating the protein PDB. The verified BT PDB was used for docking with the specified ligand. Based on an improved docking score compared to alpha-mangostin (AM), two modified compounds were identified: 1,6-dihydroxy-7-methoxy-2,8-bis(3-methylbut-2-en-1-yl)-9H-xanthen-9-one (C1) and 1,6-dihydroxy-2,8-bis(3-methylbut-2-en-1-yl)-9H-xanthen-9-one (C2). In addition, molecular dynamics simulations were conducted to analyze the interaction characteristics of the two bound BT–new compound complexes. During simulations, the TRP9, ARG50, VAL52, and GLN122 residues of BT-C1 that align to the identical residues in BT-AM generate consistent hydrogen bond interactions with 0–3 and 0–2. However, the BT-C2 complex has a different binding site, TYR 258, ILE 281, and SER 302, and can form more hydrogen bonds in the range 0–4. Therefore, this study reveals that C1 and C2 inhibit BT as an additive or synergistic effect; however, further in vitro and in vivo studies are needed.

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Section: Resultsmentioning

confidence: 99%

Homology Modeling, Molecular Docking, Molecular Dynamic Simulation, and Drug-Likeness of the Modified Alpha-Mangostin against the β-Tubulin Protein of Acanthamoeba Keratitis

et al. 2022

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“…We recently reported that psLAAO1 pre-treated with a chelating agent showed transiently increased antibacterial activity that was either decreased or diminished after one or two freeze–thaw cycles. Moreover, the mutant constructs of His348, whose position is predicted to be a metal coordination site by homology modeling simulation, did not show antibacterial activity or Mg 2+ coordination (Kasai et al 2020 (Table 1 V1)), indicating that Mg 2+ coordination is essential for the enzymatic activity and structural stability of psLAAO1 at His348. Metal detection assay revealed that pre-chelated psLAAO1 did not detect Zn 2+ , whereas 1 mole of the wild type or mutant constructs (His348Ala and His209Ala) detected approximately 0.5 mole of Zn 2+ .…”

Section: Characterization Structural Stability and Regulation By Metal Ionsmentioning

confidence: 99%

“…Further, hcLAAO4, one of the fungal LAAOs from Hebeloma cylindrosporum , was expressed in E. coli or yeast ( Pichia pastors ) systems with His-Tag (Bloess et al 2019 ; Heß et al 2020 ). Moreover, L-Lys-specific LAAO isolated from the Siganus pramin (serum) or Platichthys stellatus (gill), which is an innate immune protein with antibacterial activity, was codon optimized for expression in insect cells (Sf9) or P. pastoris expression systems (Han et al 2020 , Kasai et al 2020 (Table 1 V1)). These recombinant LAAOs can be easily high mass-produced by scale-up and display strong bioactivity via bactericidal, parasiticidal, and apoptotic effects, and bind to the surface of bacteria, similar to native LAAOs, suggesting potential use of LAAOs against clinical pathogens.…”

Section: Perspectives For Possible Biomedical Applicationsmentioning

confidence: 99%

Antimicrobial properties of L-amino acid oxidase: biochemical features and biomedical applications

Kasai

Nakano

Ohishi

et al. 2021

Appl Microbiol Biotechnol

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Mucus layer that covers the body surface of various animal functions as a defense barrier against microbes, environmental xenobiotics, and predators. Previous studies have reported that L-amino acid oxidase (LAAO), present in several animal fluids, has potent properties against pathogenic bacteria, viruses, and parasites. LAAO catalyzes the oxidative deamination of specific L-amino acids with the generation of hydrogen peroxide and L-amino acid metabolites. Further, the generated hydrogen peroxide is involved in oxidation (direct effect) while the metabolites activate immune responses (indirect effect). Therefore, LAAO exhibits two different mechanisms of bioactivation. Previously, we described the selective, specific, and local oxidative and potent antibacterial actions of various LAAOs as potential therapeutic strategies. In this review, we focus on their biochemical features, enzymatic regulations, and biomedical applications with a view of describing their probable role as biochemical agents and biomarkers for microbial infections, cancer, and autoimmune-mediated diseases. We consider that LAAOs hold implications in biomedicine owing to their antimicrobial activity wherein they can be used in treatment of infectious diseases and as diagnostic biomarkers in the above-mentioned diseased conditions. Key points •Focus on biochemical features, enzymatic regulation, and biomedical applications of LAAOs. •Mechanisms of antimicrobial activity, inflammatory regulation, and immune responses of LAAOs. •Potential biomedical application as an antimicrobial and anti-infection agent, and disease biomarker.

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Improvement of Heterologous Soluble Expression of L‐amino Acid Oxidase Using Logistic Regression

Nakahara,

Su,

Wakayama

et al. 2024

ChemBioChem

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Successful implementation of enzymes in practical application hinges on the development of efficient mass production techniques. However, in a heterologous expression system, the protein is often unable to fold correctly and, thus, forms inclusion bodies, resulting in the loss of its original activity. In this study, we present a new and more accurate model for predicting amino acids associated with an increased L‐amino acid oxidase (LAO) solubility. Expressing LAO from Rhizoctonia solani in Escherichia coli and combining random mutagenesis and statistical logistic regression, we modified 108 amino acid residues by substituting hydrophobic amino acids with serine and hydrophilic amino acids with alanine. Our results indicated that specific mutations in Euclidean distance, glycine, methionine, and secondary structure increased LAO expression. Furthermore, repeated mutations were performed for LAO based on logistic regression models. The mutated LAO displayed a significantly increased solubility, with the 6‐point and 58‐point mutants showing a 2.64‐ and 4.22‐fold increase, respectively, compared with WT‐LAO. Ultimately, using recombinant LAO in the biotransformation of α‐keto acids indicates its great potential as a biocatalyst in industrial production.

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Metal coordination by L-amino acid oxidase derived from flounder Platichthys stellatus is structurally essential and regulates antibacterial activity

Cited by 5 publications

References 33 publications

Homology Modeling, Molecular Docking, Molecular Dynamic Simulation, and Drug-Likeness of the Modified Alpha-Mangostin against the β-Tubulin Protein of Acanthamoeba Keratitis

Homology Modeling, Molecular Docking, Molecular Dynamic Simulation, and Drug-Likeness of the Modified Alpha-Mangostin against the β-Tubulin Protein of Acanthamoeba Keratitis

Antimicrobial properties of L-amino acid oxidase: biochemical features and biomedical applications

Improvement of Heterologous Soluble Expression of L‐amino Acid Oxidase Using Logistic Regression

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