scholarly journals Improvement of the Glutaryl-7-Aminocephalosporanic Acid Acylase Activity of a Bacterial γ-Glutamyltranspeptidase

2008 ◽  
Vol 74 (11) ◽  
pp. 3400-3409 ◽  
Author(s):  
Chiaki Yamada ◽  
Kyoko Kijima ◽  
Sayaka Ishihara ◽  
Chinatsu Miwa ◽  
Kei Wada ◽  
...  
Keyword(s):  
Amino Acid ◽  
Screening Method ◽  
Three Dimensional ◽  
Catalytic Efficiency ◽  
Industrial Applications ◽  
Single Amino Acid ◽  
Dimensional Structure ◽  
Acid Oxidase ◽  
Amino Acid Oxidase ◽  
K 12

ABSTRACT 7-Aminocephalosporanic acid (7-ACA) is an important material in the production of semisynthetic cephalosporins, which are the best-selling antibiotics worldwide. 7-ACA is produced from cephalosporin C via glutaryl-7-ACA (GL-7-ACA) by a bioconversion process using d-amino acid oxidase and cephalosporin acylase (or GL-7-ACA acylase). Previous studies demonstrated that a single amino acid substitution, D433N, provided GL-7-ACA acylase activity for γ-glutamyltranspeptidase (GGT) of Escherichia coli K-12. In this study, based on its three-dimensional structure, residues involved in substrate recognition of E. coli GGT were rationally mutagenized, and effective mutations were then combined. A novel screening method, activity staining followed by a GL-7-ACA acylase assay with whole cells, was developed, and it enabled us to obtain mutant enzymes with enhanced GL-7-ACA acylase activity. The best mutant enzyme for catalytic efficiency, with a k cat/Km value for GL-7-ACA almost 50-fold higher than that of the D433N enzyme, has three amino acid substitutions: D433N, Y444A, and G484A. We also suggest that GGT from Bacillus subtilis 168 can be another source of GL-7-ACA acylase for industrial applications.

Three-Dimensional Structure of Porcine Kidney D-Amino Acid Oxidase at 3.0 A Resolution

1996 ◽  
Vol 120 (1) ◽  
pp. 14-17 ◽  
Author(s):  
H. Mizutani ◽  
I. Miyahara ◽  
K. Hirotsu ◽  
Y. Nishina ◽  
K. Shiga ◽  
...  
Keyword(s):  
Amino Acid ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Acid Oxidase ◽  
Porcine Kidney ◽  
Amino Acid Oxidase ◽  
Three Dimensional Structure

P219L substitution in human D-amino acid oxidase impacts the ligand binding and catalytic efficiency

2020 ◽  
Vol 168 (5) ◽  
pp. 557-567
Author(s):  
Wanitcha Rachadech ◽  
Yusuke Kato ◽  
Rabab M Abou El-Magd ◽  
Yuji Shishido ◽  
Soo Hyeon Kim ◽  
...  
Keyword(s):  
Amino Acid ◽  
Conformational Changes ◽  
Active Site ◽  
Structural Changes ◽  
Catalytic Efficiency ◽  
Acid Oxidase ◽  
Wild Type ◽  
Amino Acid Oxidase ◽  
Adenine Ring ◽  
The Impact

Abstract Human D-amino acid oxidase (DAO) is a flavoenzyme that is implicated in neurodegenerative diseases. We investigated the impact of replacement of proline with leucine at Position 219 (P219L) in the active site lid of human DAO on the structural and enzymatic properties, because porcine DAO contains leucine at the corresponding position. The turnover numbers (kcat) of P219L were unchanged, but its Km values decreased compared with wild-type, leading to an increase in the catalytic efficiency (kcat/Km). Moreover, benzoate inhibits P219L with lower Ki value (0.7–0.9 µM) compared with wild-type (1.2–2.0 µM). Crystal structure of P219L in complex with flavin adenine dinucleotide (FAD) and benzoate at 2.25 Å resolution displayed conformational changes of the active site and lid. The distances between the H-bond-forming atoms of arginine 283 and benzoate and the relative position between the aromatic rings of tyrosine 224 and benzoate were changed in the P219L complex. Taken together, the P219L substitution leads to an increase in the catalytic efficiency and binding affinity for substrates/inhibitors due to these structural changes. Furthermore, an acetic acid was located near the adenine ring of FAD in the P219L complex. This study provides new insights into the structure–function relationship of human DAO.

scholarly journals The Molecules and Mechanisms Underlying the Antimicrobial Activity of Escapin, an L–Amino Acid Oxidase from the Ink of Sea Hares

2018 ◽  
Author(s):  
Charles D. Derby ◽  
Eric S. Gilbert ◽  
Phang C. Tai
Keyword(s):  
Antimicrobial Activity ◽  
Amino Acid ◽  
Industrial Applications ◽  
Chemical Defenses ◽  
Acid Oxidase ◽  
Marine Animals ◽  
Amino Acid Oxidase ◽  
Health And Disease ◽  
S Oxidation ◽  
Related Proteins

Many marine animals use chemicals to defend themselves and their eggs from predators. Beyond their ecologically relevant functions, these chemicals may also have properties that make them beneficial for humans, including with biomedical and industrial applications. For example, some chemical defenses are also powerful antimicrobial or anti-tumor agents with relevance to human health and disease. One such chemical defense, Escapin, an L–amino acid oxidase in the defensive ink of the sea hare Aplysia californica, and related proteins have been investigated for their biomedical properties. This review details our current understanding of Escapin’s antimicrobial activity, including the array of chemicals generated by Escapin’s oxidation of its major substrates, L–lysine and L–arginine, and mechanisms underlying these molecules’ bactericidal and bacteriostatic effects on planktonic cells and the prevention of formation and removal of bacterial biofilms. Models of Escapin’s effects are presented, and future directions are proposed.

scholarly journals Immobilization of Genetically-Modified d-Amino Acid Oxidase and Catalase on Carbon Nanotubes to Improve the Catalytic Efficiency

Catalysts ◽  
2016 ◽  
Vol 6 (5) ◽  
pp. 66 ◽  
Author(s):  
Rong Li ◽  
Jian Sun ◽  
Yaqi Fu ◽  
Kun Du ◽  
Mengsha Cai ◽  
...  
Keyword(s):  
Carbon Nanotubes ◽  
Amino Acid ◽  
Genetically Modified ◽  
Catalytic Efficiency ◽  
Acid Oxidase ◽  
Amino Acid Oxidase

Optimization of medium and cultivation conditions forl-amino acid oxidase production byAspergillus fumigatus

2009 ◽  
Vol 55 (9) ◽  
pp. 1096-1102 ◽  
Author(s):  
Susmita Singh ◽  
B. K. Gogoi ◽  
R. L. Bezbaruah
Keyword(s):  
Enzyme Activity ◽  
Amino Acid ◽  
Cell Mass ◽  
Industrial Applications ◽  
High Yield ◽  
Acid Oxidase ◽  
Amino Acid Oxidase ◽  
North East ◽  
The North ◽  
Dry Cell

A fungal strain was selected from the microbial repository of the North-East Institute of Science and Technology, Jorhat, India, which could produce a high yield of l-amino acid oxidase. 18SrRNA, ITS1, 5.8SrRNA ITS2, and partial 28 S rRNA sequencing and phenotypic characteristics indicate that it belong to the species Aspergillus fumigatus (designated as P13). Maximum production of enzyme (59.55 × 10−3 U/mg dry cell mass) was obtained in a medium containing 10 g/L glucose, 4 g/L yeast extract, and 4 g/L ammonium sulfate, with 20 mmol/L of l-threonine as the inducer. The optimum temperature for enzyme production was 30 °C at pH 7.0, with a shaking speed of 200 r/min. At 96 h, the enzyme activity was maximum. The A. fumigatus P13 l-amino acid oxidase accepts a broad substrate range, and the maximum enzyme activity (20.41 × 10−3 U/mg dry cell mass) was obtained with 50 mmol/L of dl-tyrosine. In the literature, no reports have been found regarding the production of l-amino acid oxidase by A. fumigatus. The enzyme showed enantiomerically pure amino acid formation, which has tremendous demand in industrial applications.

Mutant d-amino acid oxidase with higher catalytic efficiency toward d-amino acids with bulky side chains

2012 ◽  
Vol 61 (7) ◽  
pp. 1489-1496 ◽  
Author(s):  
N. V. Komarova ◽  
I. V. Golubev ◽  
S. V. Khoronenkova ◽  
V. I. Tishkov
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Catalytic Efficiency ◽  
Side Chains ◽  
Acid Oxidase ◽  
Amino Acid Oxidase

Purification and Characterization of Lupus Anticoagulant Like Protein from Agkistrodon Halys Brevicaudus Venom

1996 ◽  
Vol 76 (06) ◽  
pp. 0993-0997
Author(s):  
Zhao-Yan Li ◽  
Xiao-Wei Wu ◽  
Tie-Fu Yu ◽  
Eric C-Y Lian
Keyword(s):  
Amino Acid ◽  
Lupus Anticoagulant ◽  
Inhibitory Effect ◽  
Clotting Factor ◽  
Acid Oxidase ◽  
Crude Venom ◽  
Amino Acid Oxidase ◽  
Sds Page ◽  
The Individual ◽  
Reducing Condition

SummaryBy means of CM-Sephadex C-25, DEAE-Sephadex A-50, Sephadex G-200, and Sephadex G-75 chromatographies, a lupus anticoagulant like protein (LALP) from Agkistrodon halys brevicaudus was purified. On SDS-PAGE, the purified LALP had a molecular weight of 25,500 daltons under non-reducing condition and 15,000 daltons under reducing condition. The isoelectric point was pH 5.6. Its N terminal amino acid sequencing revealed a mixture of 2 sequences: DCP(P/S)(D/G)WSSYEGH(C/R)Q(Q/K). It was devoid of phospho-lipaseA, fibrino(geno)lytic, 5′-nucleotidase, L-amino acid oxidase, phosphomonoesterase, phosphodiesterase and thrombin-like activities, which were found in crude venom. In the presence of LALP, PT, aPTT, and dRVVT of human plasma were markedly prolonged and its effects were concentration-dependent but time-independent. The inhibitory effect of LALP on the plasma clotting time was enhanced by decreasing phospholipid concentration in TTI test. The individual clotting factor activity was not affected by LALP when higher dilutions of LALP-plasma mixture were used for assay. Russell’s viper venom time was shortened when high phospholipid confirmatory reagent was used. Therefore, the protein has lupus anticoagulant property.

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