scholarly journals Physical-Chemical Properties of L-asparaginase Mutants From Rhodospirillum Rubrum which Showed Antitelomerase Activity

2019 ◽  
Vol 2 (1) ◽  
Author(s):  
M.V. Pokrovskaya ◽  
S.S. Aleksandrova ◽  
A.V. Veselovsky ◽  
D.D. Zdanov ◽  
V.S. Pokrovsky ◽  
...  
Keyword(s):  
Amino Acid ◽  
Rhodospirillum Rubrum ◽  
Chemical Properties ◽  
Telomerase Activity ◽  
Kinetic Properties ◽  
Ph Optimum ◽  
Physical Chemical ◽  
Asparaginase Activity ◽  
Terminal Amino ◽  
Mutant Forms

Rru_A3730 protein is a bacterial Rhodospirillum rubrum L-asparaginase (RrA), which is known by its anticancer activity. RrA variants with point amino acid substitutions in the region of 150 amino acids residues: RrA17N, K149E, RrAE149R, V150P, F151T, RrА17N, E149R, V150P, RrAE149R, V150P, showed antiproliferative properties, and also by their ability to suppress telomerase activity. This work is devoted to comparison of physical-chemical and catalytic properties of these mutant forms of RrA. It is shown that pH optimum is in the alkaline zone (8.5 – 9.3); L-glutaminase and D-asparaginase activity is respectively not more than 0.1% and 1.6% of L-asparaginase for all studied variants of RrA. The presence of the N17-terminal amino acid sequence MASMTGGQMGRGSSRQ of the capsid protein of bacteriophage T7 in the RrA structure leads to an increase in the thermal stability of mutant RrA analogues (from 50°C to 56°C) and their resistance to denaturation in the presence of 3 – 4 M urea. It is of Metal ions exhibit multidirectional effects on L-asparaginase activity of RrA. K+, Ca2+, Zn2+, Cs+, Co2+ in significantly affect the activity of L-asparaginase, while Mn2+, Cu2+, Fe3+ ions inhibit it. There was no correlation between antitelomerase (antiproliferative) activity and kinetic properties of mutant forms of L-asparaginase RrA.

Physicochemical properties of a novel α-L-arabinofuranosidase fromRhizomucor pusillusHHT-1

2001 ◽  
Vol 47 (8) ◽  
pp. 767-772 ◽  
Author(s):  
A KM Shofiqur Rahman ◽  
Shinya Kawamura ◽  
Masahiro Hatsu ◽  
M M Hoq ◽  
Kazuhiro Takamizawa
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Molecular Mass ◽  
Gel Filtration ◽  
Hydrolytic Activity ◽  
Exchange Chromatography ◽  
Ph Optimum ◽  
Rhizomucor Pusillus ◽  
Terminal Amino ◽  
Metal Cofactors

The zygomycete fungus Rhizomucor pusillus HHT-1, cultured on L(+)arabinose as a sole carbon source, produced extracellular α-L-arabinofuranosidase. The enzyme was purified by (NH4)2SO4fractionation, gel filtration, and ion exchange chromatography. The molecular mass of this monomeric enzyme was 88 kDa. The native enzyme had a pI of 4.2 and displayed a pH optimum and stability of 4.0 and 7.0–10.0, respectively. The temperature optimum was 65°C, and it was stable up to 70°C. The Kmand Vmaxfor p-nitrophenyl α-L-arabinofuranoside were 0.59 mM and 387 µmol·min–1·mg–1protein, respectively. Activity was not stimulated by metal cofactors. The N-terminal amino acid sequence did not show any similarity to other arabinofuranosidases. Higher hydrolytic activity was recorded with p-nitrophenyl α-L-arabinofuranoside, arabinotriose, and sugar beet arabinan; lower hydrolytic activity was recorded with oat–spelt xylan and arabinogalactan, indicating specificity for the low molecular mass L(+)-arabinose containing oligosaccharides with furanoside configuration.Key words: α-L-arabinofuranosidase, enzyme purification, amino acid sequence, Rhizomucor pusillus.

scholarly journals Physical-chemical properties of C-phycocyanin isolated from an acido-thermophilic eukaryote, Cyanidium caldarium

1975 ◽  
Vol 147 (1) ◽  
pp. 63-70 ◽  
Author(s):  
O H Kao ◽  
M R Edwards ◽  
D S Berns
Keyword(s):  
Absorption Spectrum ◽  
High Temperature ◽  
Amino Acid ◽  
Ph Dependence ◽  
Chemical Properties ◽  
Sedimentation Velocity ◽  
Subunit Structure ◽  
Cyanidium Caldarium ◽  
Eukaryotic Alga ◽  
Physical Chemical

C-Phycocyanin from an acido-thermophilic eukaryotic alga, Cyanidium caldarium, was characterized with respect to subunit structure, absorption spectrum and fluorescence properties and was found to be similar to C-phycocyanins from mesophilic sources. The pH-dependence of fluorescence polarization and the changes in sedimentation velocity as a function of pH, concentration and temperature indicate the presence of extremely large amounts of unusually stable 19S aggregates. It was not possible to disaggregate this phycocyanin completely to monomer under normal conditions. The amino acid composition is similar to that of phycocyanins from other thermophilic and halophilic sources. The isoelectric point of this C-phycocyanin was 5.11, an unusually high value. The properties of this C-phycocyanin suggest an increase in protein stability as its mode of adaptation to the environmental stress of high temperature.

Properties of Purified Quinonoid Dihydropterin Reductase

1973 ◽  
Vol 51 (9) ◽  
pp. 1229-1239 ◽  
Author(s):  
Surinder Cheema ◽  
S. J. Soldin ◽  
Antoinetta Knapp ◽  
T. Hofmann ◽  
K. G. Scrimgeour
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Adrenal Medulla ◽  
Kinetic Properties ◽  
Terminal Amino Acid ◽  
Sheep Liver ◽  
Wide Range ◽  
Sheep Brain ◽  
Terminal Amino ◽  
Covalently Linked

Quinonoid dihydropterin reductase has been purified to homogeneity from sheep liver, sheep brain, and beef adrenal medulla. Each of these enzymes has a molecular weight of about 45 000–55 000, and is composed of two subunits of half that weight. The subunits of the sheep liver reductase have identical charge, size, and N-terminal amino acid residue. The reductase exists in solution over a wide range of concentrations as the dimer. A dimer covalently linked by dimethylsuberimidate retains full activity. A number of kinetic properties of quinonoid dihydropterin reductase, including inhibition by thiol reagents and by pterin analogues, are reported.

scholarly journals Relationship between Metabolic Fluxes and Sequence-Derived Properties of Enzymes

2014 ◽  
Vol 2014 ◽  
pp. 1-9 ◽  
Author(s):  
Peteris Zikmanis ◽  
Inara Kampenusa
Keyword(s):  
Metabolic Pathways ◽  
Chemical Properties ◽  
Structural Features ◽  
Kinetic Properties ◽  
Net Charge ◽  
Metabolic Fluxes ◽  
Physical Chemical ◽  
Glycolysis Pathway ◽  
Derived Properties ◽  
Linear Regressions

Metabolic fluxes are key parameters of metabolic pathways being closely related to the kinetic properties of enzymes, thereby could be dependent on. This study examines possible relationships between the metabolic fluxes and the physical-chemical/structural features of enzymes from the yeast Saccharomyces cerevisiae glycolysis pathway. Metabolic fluxes were quantified by the COPASI tool using the kinetic models of Hynne and Teusink at varied concentrations of external glucose. The enzyme sequences were taken from the UniProtKB and the average amino acid (AA) properties were computed using the set of Georgiev’s uncorrelated scales that satisfy the VARIMAX criterion and specific AA indices that show the highest correlations with those. Multiple linear regressions (88.41% < Radjusted2< 93.32%; P<0.00001) were found between the values of metabolic fluxes and the selected sets of the average AA properties. The hydrophobicity, α-helicity, and net charge were pointed out as the most influential characteristics of the sequences. The results provide an evidence that metabolic fluxes of the yeast glycolysis pathway are closely related to certain physical-chemical properties of relevant enzymes and support the view on the interdependence of catalytic, binding, and structural AA residues to ensure the efficiency of biocatalysts and, hence, physiologically adequate metabolic processes.

scholarly journals Studies on the coagulant enzyme from Agkistrodon rhodostoma venom. Isolation and some properties of the enzyme

1973 ◽  
Vol 131 (4) ◽  
pp. 799-807 ◽  
Author(s):  
M. W. C. Hatton
Keyword(s):  
Amino Acid ◽  
Esterase Activity ◽  
Polypeptide Chain ◽  
Kinetic Properties ◽  
Amino Acid Residues ◽  
Commercial Preparation ◽  
Single Polypeptide Chain ◽  
Coagulant Activity ◽  
Terminal Amino ◽  
Single Polypeptide

1. Arvin, a commercial preparation of the coagulant activity from the venom of Agkistrodon rhodostoma, is shown to contain a non-coagulant caseinolytic fraction. 2. A method is described for the purification of the coagulant enzyme free from any detectable contaminating protein. 3. The coagulant enzyme is identified as a glycoprotein which probably consists of a single polypeptide chain containing approx. 29% by weight of carbohydrate. Amino acid and carbohydrate analyses are reported and the N- and C-terminal amino acid residues identified. 4. Electrophoresis on polyacrylamide gel reveals the polymorphic nature of the glycoprotein. Five forms of the enzyme are observed. 5. The coagulant action is correlated with an arginine esterase activity and kinetic properties are studied with both arginine and lysine esters as substrates. The inhibitory nature of guanidine and arginine toward the esterase activity is reported.

scholarly journals High sequence homology between protein tyrosine acid phosphatase from boar seminal vesicles and human prostatic acid phosphatase.

2009 ◽  
Vol 56 (3) ◽  
Author(s):  
Paweł Wysocki ◽  
Grazyna Płucienniczak ◽  
Jerzy Strzezek
Keyword(s):  
Acid Phosphatase ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Seminal Vesicle ◽  
Prostatic Acid Phosphatase ◽  
Kinetic Properties ◽  
Terminal Amino Acid ◽  
Protein Tyrosine ◽  
Terminal Amino ◽  
Terminal Amino Acid Sequence

Boar seminal vesicle protein tyrosine acid phosphatase (PTAP) and human prostatic acid phosphatase (PAP) show high affinity for protein phosphotyrosine residues. The physico-chemical and kinetic properties of the boar and human enzymes are different. The main objective of this study was to establish the nucleotide sequence of cDNA encoding boar PTAP and compare it with that of human PAP cDNA. Also, the amino-acid sequence of boar PTAP was compared with the sequence of human PAP. PTAP was isolated from boar seminal vesicle fluid and sequenced. cDNA to boar seminal vesicle RNA was synthesized, amplified by PCR, cloned in E. coli and sequenced. The obtained N-terminal amino-acid sequence of boar PTAP showed 92% identity with the N-terminal amino-acid sequence of human PAP. The determined sequence of a 354 bp nucleotide fragment (GenBank accession number: GQ184596) showed 90% identity with the corresponding sequence of human PAP. On the basis of this sequence a 118 amino acid fragment of boar PTAP was predicted. This fragment showed 89% identity with the corresponding fragment of human PAP and had a similar hydropathy profile. The compared sequences differ in terms of their isoelectric points and amino-acid composition. This may explain the differences in substrate specificity and inhibitor resistance of boar PTAP and human PAP.

scholarly journals Suppression of telomerase activity leukemic cells by mutant forms of Rhodospirillum rubrum L-asparaginase

2017 ◽  
Vol 63 (1) ◽  
pp. 62-74 ◽  
Author(s):  
M.V. Pokrovskaya ◽  
D.D. Zhdanov ◽  
M.A. Eldarov ◽  
S.S. Aleksandrova ◽  
A.V. Veselovskiy ◽  
...  
Keyword(s):  
Rhodospirillum Rubrum ◽  
Erwinia Carotovora ◽  
Telomerase Activity ◽  
Jurkat Cells ◽  
Site Directed Mutagenesis ◽  
Leukemic Cells ◽  
Type I ◽  
Wolinella Succinogenes ◽  
Stable Mutant ◽  
Mutant Forms

The active and stable mutant forms of short chain cytoplasmic L-asparaginase type I of Rhodospirillum rubrum (RrA): RrA+N17, D60K, F61L, RrA+N17, A64V, E67K, RrA+N17, E149R, V150P, RrAE149R, V150P and RrAE149R, V150P, F151T were obtained by the method of site-directed mutagenesis. It is established that variants RrA-N17, E149R, V150P, F151T and RrАE149R, V150P are capable to reduce an expression hTERT subunit of telomerase and, hence, activity of telomeres in Jurkat cells, but not in cellular lysates. During too time, L-asparaginasеs of Escherichia coli, Erwinia carotovora and Wolinella succinogenes, mutant forms RrА+N17, D60K, F61L and RrА+N17, A64V, E67K do not suppress of telomerase activity. The assumption of existence in structure RrA of areas (amino acids residues in the position 146-164, 1-17, 60-67) which are responsible for suppression of telomerase activity is made. The received results show that antineoplastic activity of some variants RrA is connected both with reduction of concentration of free L-asparagine, and with expression suppression of hTERT telomerase subunit, that opens new prospects for antineoplastic therapy.

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