Ancestral L-amino acid oxidases for deracemization and stereoinversion of amino acids

Nakano, Shogo; Kozuka, Kohei; Minamino, Yuki; Karasuda, H.; Hasebe, Fumihito; Ito, Sohei

doi:10.1038/s42004-020-00432-8

Cited by 30 publications

(45 citation statements)

References 55 publications

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“…by Blastp utilizing open reading frames of six Pseudoalteromonas genera as a library. As shown in a previous study, these genera have LAAO sequences that were utilized to design AncLAAO-N5 (26,28). Through this analysis, a paralog sequence was assigned in Pseudoalteromonas luteoviolacea; the sequence identity was quite low compared with the functionally annotated LAAOs.…”

Section: Sequence Classification From Databases To Assign a New Llyso Familymentioning

confidence: 85%

“…Currently, four enzymes which belong to the FAO superfamily and exhibit LAAO activity are reported. These include AncLAAO (26,28), VioA (31), AROD (27,30), and AncLLysO. These enzymes share low sequence identity to each other and to other proteins in the superfamily registered in the PDB database (<30%).…”

Section: J O U R N a L P R E -P R O O Fmentioning

confidence: 99%

“…Therefore, ASR is currently being adopted as a tool for protein engineering (25). For example, new LAAOs which bear broad substrate selectivity (> 10 Lamino acids) could be assigned by a paralog search of L-arginine oxidase (AROD) (26)(27)(28). Artificial LAAOs can be designed by ASR utilizing six of the assigned LAAOs as a sequence library.…”

Section: Introductionmentioning

confidence: 99%

“…On the other hand, there are thousands of protein sequences belonging to the LAAO superfamily in databases (29), and the currently available functionally characterized sequences are only the tip of the iceberg. Several sequences, such as AROD (27,30), AncLAAO (26,28), and Ltryptophan oxidase (VioA) (31)(32)(33), can now be functionally annotated. Thus, there is an opportunity to acquire new LAAOs from the database using in silico enzyme screening methods.…”

Section: Introductionmentioning

confidence: 99%

“…In this study, we attempted to assign new LAAOs from the database utilizing one of the previously designed AncLAAO sequences (28) as a template. Through combinational approaches using paralog searches, the application of previously reported original protein sequence selection methods (27,(34)(35)(36), and biochemical assays, we newly characterized LAAOs exhibiting high specificity toward L-Lys.…”

Section: Introductionmentioning

confidence: 99%

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Catalytic mechanism of ancestral L-lysine oxidase assigned by sequence data mining

Sugiura

Nakano

Niwa

et al. 2021

Journal of Biological Chemistry

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Section: Sequence Classification From Databases To Assign a New Llyso Familymentioning

confidence: 85%

Section: J O U R N a L P R E -P R O O Fmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Catalytic mechanism of ancestral L-lysine oxidase assigned by sequence data mining

Sugiura

Nakano

Niwa

et al. 2021

Journal of Biological Chemistry

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Crystal structure and enzyme engineering of the broad substrate spectrum l‐amino acid oxidase 4 from the fungus Hebeloma cylindrosporum

Koopmeiners,

Gilzer,

Widmann

et al. 2024

FEBS Letters

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l‐Amino acid oxidases (LAAOs) catalyze the oxidative deamination of l‐amino acids to α‐keto acids. Recombinant production of LAAOs with broad substrate spectrum remains a formidable challenge. We previously achieved this for the highly active and thermostable LAAO4 of Hebeloma cylindrosporum (HcLAAO4). Here, we crystallized a proteolytically truncated surface entropy reduction variant of HcLAAO4 and solved its structure in substrate‐free form and in complex with diverse substrates. The ability to support the aliphatic portion of a substrate's side chain by an overall hydrophobic active site is responsible for the broad substrate spectrum of HcLAAO4, including l‐amino acids with big aromatic, acidic and basic side chains. Based on the structural findings, we generated an E288H variant with increased activity toward pharmaceutical building blocks of high interest.

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Chemoenzymatic Production of Enantiocomplementary 2‐Substituted 3‐Hydroxycarboxylic Acids from l‐α‐Amino Acids

et al. 2021

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A two‐enzyme cascade reaction plus in situ oxidative decarboxylation for the transformation of readily available canonical and non‐canonical l‐α‐amino acids into 2‐substituted 3‐hydroxycarboxylic acid derivatives is described. The biocatalytic cascade consisted of an oxidative deamination of l‐α‐amino acids by an l‐α‐amino acid deaminase from Cosenzaea myxofaciens, rendering 2‐oxoacid intermediates, with an ensuing aldol addition reaction to formaldehyde, catalyzed by metal‐dependent (R)‐ or (S)‐selective carboligases namely 2‐oxo‐3‐deoxy‐l‐rhamnonate aldolase (YfaU) and ketopantoate hydroxymethyltransferase (KPHMT), respectively, furnishing 3‐substituted 4‐hydroxy‐2‐oxoacids. The overall substrate conversion was optimized by balancing biocatalyst loading and amino acid and formaldehyde concentrations, yielding 36–98% aldol adduct formation and 91–98% ee for each enantiomer. Subsequent in situ follow‐up chemistry via hydrogen peroxide‐driven oxidative decarboxylation afforded the corresponding 2‐substituted 3‐hydroxycarboxylic acid derivatives.

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Ancestral L-amino acid oxidases for deracemization and stereoinversion of amino acids

Cited by 30 publications

References 55 publications

Catalytic mechanism of ancestral L-lysine oxidase assigned by sequence data mining

Catalytic mechanism of ancestral L-lysine oxidase assigned by sequence data mining

Crystal structure and enzyme engineering of the broad substrate spectrum l‐amino acid oxidase 4 from the fungus Hebeloma cylindrosporum

Chemoenzymatic Production of Enantiocomplementary 2‐Substituted 3‐Hydroxycarboxylic Acids from l‐α‐Amino Acids

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