scholarly journals Contribution of the dimeric state to the thermal stability of the flavoprotein D-amino acid oxidase

2003 ◽  
Vol 12 (5) ◽  
pp. 1018-1029 ◽  
Author(s):  
Loredano Pollegioni ◽  
Stefania Iametti ◽  
Dimitrios Fessas ◽  
Laura Caldinelli ◽  
Luciano Piubelli ◽  
...  
Keyword(s):  
Thermal Stability ◽  
Amino Acid ◽  
Acid Oxidase ◽  
Amino Acid Oxidase ◽  
Thermal Stability Of

scholarly journals Study of the Structure-Function-Stability Relationships in Yeast D-amino Acid Oxidase: Hydrophobization of Alpha-Helices

Acta Naturae ◽  
2014 ◽  
Vol 6 (3) ◽  
pp. 76-88 ◽  
Author(s):  
I. V. Golubev ◽  
N. V. Komarova ◽  
K. V. Ryzhenkova ◽  
T. A. Chubar ◽  
S. S. Savin ◽  
...  
Keyword(s):  
Thermal Stability ◽  
Amino Acid ◽  
3D Structure ◽  
Amino Acid Sequences ◽  
Acid Oxidase ◽  
Wild Type ◽  
Wild Type Enzyme ◽  
Amino Acid Oxidase ◽  
Thermal Stabilities ◽  
Alpha Helices

Hydrophobization of alpha-helices is one of the general approaches used for improving the thermal stability of enzymes. A total of 11 serine residues located in alpha-helices have been found based on multiple alignments of the amino acid sequences of D-amino acid oxidases from different organisms and the analysis of the 3D-structure of D-amino acid oxidase from yeast Trigonopsis variabilis (TvDAAO, EC 1.4.3.3). As a result of further structural analysis, eight Ser residues in 67, 77, 78, 105, 270, 277, 335, and 336 positions have been selected to be substituted with Ala. S78A and S270A substitutions have resulted in dramatic destabilization of the enzyme. Mutant enzymes were inactivated during isolation from cells. Another six mutant TvDAAOs have been highly purified and their properties have been characterized. The amino acid substitutions S277A and S336A destabilized the protein globule. The thermal stabilities of TvDAAO S77A and TvDAAO S335A mutants were close to that of the wild-type enzyme, while S67A and S105A substitutions resulted in approximately 1.5- and 2.0-fold increases in the TvDAAO mutant thermal stability, respectively. Furthermore, the TvDAAO S105A mutant showed on average a 1.2- to 3.0-fold higher catalytic efficiency with D-Asn, D-Tyr, D-Phe, and D-Leu as compared to the wild-type enzyme.

scholarly journals X-ray structure analysis of a unique D-amino-acid oxidase from the thermophilic fungus Rasamsonia emersonii strain YA

2020 ◽  
Vol 76 (11) ◽  
pp. 517-523
Author(s):  
Yuya Shimekake ◽  
Yuki Hirato ◽  
Rikako Funabashi ◽  
Sayoko Okazaki ◽  
Masaru Goto ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Thermal Stability ◽  
Amino Acids ◽  
Amino Acid ◽  
Disulfide Bond ◽  
Thermophilic Fungus ◽  
Diffusion Method ◽  
Acid Oxidase ◽  
Amino Acid Oxidase ◽  
Intramolecular Disulfide Bond

D-Amino-acid oxidases (DAAOs) catalyze the oxidative deamination of neutral and basic D-amino acids. The DAAO from the thermophilic fungus Rasamsonia emersonii strain YA (ReDAAO) has a high thermal stability and a unique broad substrate specificity that includes the acidic D-amino acid D-Glu as well as various neutral and basic D-amino acids. In this study, ReDAAO was crystallized by the hanging-drop vapor-diffusion method and its crystal structure was determined at a resolution of 2.00 Å. The crystal structure of the enzyme revealed that unlike other DAAOs, ReDAAO forms a homotetramer and contains an intramolecular disulfide bond (Cys230–Cys285), suggesting that this disulfide bond is involved in the higher thermal stability of ReDAAO. Moreover, the structure of the active site and its vicinity in ReDAAO indicates that Arg97, Lys99, Lys114 and Ser231 are candidates for recognizing the side chain of D-Glu.

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.

Impairment of Platelet Aggregation by Echis colorata Venom Mediated by L-Amino Acid Oxidase or H2O2

1982 ◽  
Vol 48 (03) ◽  
pp. 277-282 ◽  
Author(s):  
I Nathan ◽  
A Dvilansky ◽  
T Yirmiyahu ◽  
M Aharon ◽  
A Livne
Keyword(s):  
Hydrogen Peroxide ◽  
Amino Acid ◽  
Platelet Aggregation ◽  
Snake Venom ◽  
Oxidase Activity ◽  
Enzyme Preparation ◽  
Acid Oxidase ◽  
Crude Venom ◽  
Amino Acid Oxidase ◽  
Hemostatic Disorders

SummaryEchis colorata bites cause impairment of platelet aggregation and hemostatic disorders. The mechanism by which the snake venom inhibits platelet aggregation was studied. Upon fractionation, aggregation impairment activity and L-amino acid oxidase activity were similarly separated from the crude venom, unlike other venom enzymes. Preparations of L-amino acid oxidase from E.colorata and from Crotalus adamanteus replaced effectively the crude E.colorata venom in impairment of platelet aggregation. Furthermore, different treatments known to inhibit L-amino acid oxidase reduced in parallel the oxidase activity and the impairment potency of both the venom and the enzyme preparation. H2O2 mimicked characteristically the impairment effects of L-amino acid oxidase and the venom. Catalase completely abolished the impairment effects of the enzyme and the venom. It is concluded that hydrogen peroxide formed by the venom L-amino acid oxidase plays a role in affecting platelet aggregation and thus could contribute to the extended bleeding typical to persons bitten by E.colorata.

scholarly journals Spinal mechanisms contributing to the development of pain hypersensitivity induced by sphingolipids in the rat

2021 ◽  
Author(s):  
Hong Wei ◽  
Zuyue Chen ◽  
Ari Koivisto ◽  
Antti Pertovaara
Keyword(s):  
Amino Acid ◽  
Nmda Receptors ◽  
Receptor Antagonist ◽  
Gap Junction ◽  
Male Rats ◽  
Acid Oxidase ◽  
Amino Acid Oxidase ◽  
Mk 801 ◽  
Mechanical Hypersensitivity ◽  
Pain Hypersensitivity

Abstract Background Earlier studies show that endogenous sphingolipids can induce pain hypersensitivity, activation of spinal astrocytes, release of proinflammatory cytokines and activation of TRPM3 channel. Here we studied whether the development of pain hypersensitivity induced by sphingolipids in the spinal cord can be prevented by pharmacological inhibition of potential downstream mechanisms that we hypothesized to include TRPM3, σ1 and NMDA receptors, gap junctions and D-amino acid oxidase. Methods Experiments were performed in adult male rats with a chronic intrathecal catheter for spinal drug administrations. Mechanical nociception was assessed with monofilaments and heat nociception with radiant heat. N,N-dimethylsphingosine (DMS) was administered to induce pain hypersensitivity. Ononetin, isosakuranetin, naringenin (TRPM3 antagonists), BD-1047 (σ1 receptor antagonist), carbenoxolone (a gap junction decoupler), MK-801 (NMDA receptor antagonist) and AS-057278 (inhibitor of D-amino acid oxidase, DAAO) were used to prevent the DMS-induced hypersensitivity, and pregnenolone sulphate (TRPM3 agonist) to recapitulate hypersensitivity. Results DMS alone produced within 15 min a dose-related mechanical hypersensitivity that lasted at least 24 h, without effect on heat nociception. Preemptive treatments with ononetin, isosakuranetin, naringenin, BD-1047, carbenoxolone, MK-801 or AS-057278 attenuated the development of the DMS-induced hypersensitivity, but had no effects when administered alone. Pregnenolone sulphate (TRPM3 agonist) alone induced a dose-related mechanical hypersensitivity that was prevented by ononetin, isosakuranetin and naringenin. Conclusions Among spinal pronociceptive mechanisms activated by DMS are TRPM3, gap junction coupling, the σ1 and NMDA receptors, and DAAO.

scholarly journals Picosecond laser fluorometry of FAD of D-amino acid oxidase-benzoate complex.

1983 ◽  
Vol 258 (6) ◽  
pp. 3799-3802
Author(s):  
K Yagi ◽  
F Tanaka ◽  
N Nakashima ◽  
K Yoshihara
Keyword(s):  
Amino Acid ◽  
Picosecond Laser ◽  
Acid Oxidase ◽  
Amino Acid Oxidase
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