scholarly journals Purification and Characterization of Catalase from Marine BacteriumAcinetobactersp. YS0810

2014 ◽  
Vol 2014 ◽  
pp. 1-7 ◽  
Author(s):  
Xinhua Fu ◽  
Wei Wang ◽  
Jianhua Hao ◽  
Xianglin Zhu ◽  
Mi Sun
Keyword(s):  
Amino Acid ◽  
Ion Exchange ◽  
Gel Filtration ◽  
Protoporphyrin Ix ◽  
Hydroxylamine Hydrochloride ◽  
Sodium Dithionite ◽  
Flight Mass Spectrometry ◽  
Alkali Stability ◽  
High Alkali ◽  
High Degree

The catalase from marine bacteriumAcinetobactersp. YS0810 (YS0810CAT) was purified and characterized. Consecutive steps were used to achieve the purified enzyme as follows: ethanol precipitation, DEAE Sepharose ion exchange, Superdex 200 gel filtration, and Resource Q ion exchange. The active enzyme consisted of four identical subunits of 57.256 kDa. It showed a Soret peak at 405 nm, indicating the presence of iron protoporphyrin IX. The catalase was not apparently reduced by sodium dithionite but was inhibited by 3-amino-1,2,4-triazole, hydroxylamine hydrochloride, and sodium azide. Peroxidase-like activity was not found with the substrate o-phenylenediamine. So the catalase was determined to be a monofunctional catalase. N-terminal amino acid of the catalase analysis gave the sequence SQDPKKCPVTHLTTE, which showed high degree of homology with those of known catalases from bacteria. The analysis of amino acid sequence of the purified catalase by matrix-assisted laser desorption ionization time-of-flight mass spectrometry showed that it was a new catalase, in spite of its high homology with those of known catalases from other bacteria. The catalase showed high alkali stability and thermostability.

scholarly journals Isolation and a partial amino acid sequence of insulin from the islet tissue of cod (Gadus callarias)

1968 ◽  
Vol 106 (2) ◽  
pp. 531-541 ◽  
Author(s):  
P T Grant ◽  
K. B. M. Reid
Keyword(s):  
Amino Acid ◽  
Ion Exchange ◽  
Gel Filtration ◽  
Bovine Insulin ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Performic Acid ◽  
Amino Acid Residues ◽  
Islet Tissue ◽  
A Chain

1. Insulin has been isolated by gel filtration and ion-exchange chromatography from extracts of the discrete islet tissue of cod. The final preparation yielded a single band on electrophoresis at two pH values. The biological potency was 11·5 international units/mg. in mouse-convulsion and other assay procedures. 2. Glycine and methionine were shown to be the N-terminal amino acids of the A and B chains respectively. An estimate of the molecular weight together with amino acid analyses indicated that cod insulin, like the bovine hormone, consists of 51 amino acid residues. In contrast, the amino acid composition differs markedly from bovine insulin. 3. Oxidation of insulin with performic acid yielded the A and B peptide chains, which were separated by ion-exchange chromatography. Sequence studies on smaller peptides isolated from enzymic digests or from dilute acetic acid hydrolysates of the two chains have established the sequential order of 14 of the 21 amino acid residues of the A chain and 25 of the 30 amino acid residues of the B chain.

scholarly journals Studies on Marsupial Protein. II. Amino Acid Sequence of the -Chain of Haemoglobin from the Grey Kangaroo, Macropus Giganteus

1969 ◽  
Vol 22 (6) ◽  
pp. 1437 ◽  
Author(s):  
GM Air ◽  
EOP Thompson
Keyword(s):  
Amino Acid ◽  
Ion Exchange ◽  
Amino Acid Sequence ◽  
Gel Filtration ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Amino Acid Sequences ◽  
Tryptic Digestion ◽  
Grey Kangaroo

The amino acid sequence of the jS-chain of haemoglobin from M. giganteus has been determined. The soluble peptides formed by tryptic digestion were isolated by gel filtration, ion-exchange chromatography, and paper ionophoresis, and amino acid sequences determined by the "dansyl"-Edman procedure. Special procedures were necessary for three peptides which were insoluble.

PURIFICATION OF HUMAN PARATHYROID HORMONE: RECENT STUDIES AND FURTHER OBSERVATIONS

1978 ◽  
Vol 78 (1) ◽  
pp. 49-58 ◽  
Author(s):  
H. T. KEUTMANN ◽  
G. N. HENDY ◽  
M. BOEHNERT ◽  
J. L. H. O'RIORDAN ◽  
J. T. POTTS
Keyword(s):  
Parathyroid Hormone ◽  
Amino Acid ◽  
Ion Exchange ◽  
Amino Acid Composition ◽  
Acid Composition ◽  
Trichloroacetic Acid ◽  
Gel Filtration ◽  
Exchange Chromatography ◽  
Human Parathyroid Hormone ◽  
Successive Extraction

During the isolation of human parathyroid hormone there is an extensive loss of immunoassayable hormone over the successive extraction steps, due in part to the presence of fragments that are soluble in 4% trichloroacetic acid. These fragments are derived from both the amino- and carboxyl-terminal regions of the hormone. The hormonal fractions precipitated with trichloroacetic acid were further purified by gel filtration and ion-exchange chromatography. At the final ion-exchange purification step, some preparations of the hormone eluted in multiple fractions. When the various components were characterized separately by immunoassay, amino acid composition, enzymic cleavage and partial sequence analysis, they were found to be closely comparable, although the most acidic fraction contained a blocked terminal amino group. Extraction of a number of batches of tissue permitted revision of the amino acid composition of human parathyroid hormone. Biosynthetic studies with labelled amino acids confirmed the absence of tyrosine and the presence of phenylalanine and threonine and localized these residues to definite regions of the molecule.

scholarly journals Myoglobins of Cartilaginous Fishes III. Amino Acid Sequence of Myoglobin of the Shark Galeorhinus Australis

1981 ◽  
Vol 34 (1) ◽  
pp. 5 ◽  
Author(s):  
WK Fisher ◽  
DD Koureas ◽  
EOP Thompson
Keyword(s):  
Amino Acid ◽  
Ion Exchange ◽  
Amino Acid Sequence ◽  
Gel Filtration ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Red Muscle ◽  
Cartilaginous Fishes ◽  
Port Jackson

Myoglobin isolated from the red muscle of the school shark Galeorhinus australis was purified by gel filtration and ion-exchange chromatography. The amino acid sequence was determined following digestion with trypsin and purification of the peptides by paper ionophoresis and chromatography. Sequences of purified peptides were determined by the dansyl-Edman procedure and the peptides aligned by homology with the sequence of the myoglobin of the gummy shark Mustelus antarcticus. The two myoglobin sequences showed a marked similarity (16 differences), but both sequences showed approximately the same number of differences (68) from myoglobin of the Port Jackson shark Heterodontus portusjacksoni. There are 19 residues unique to the three shark myoglobin sequences.

scholarly journals A collagenous glycoprotein found in dissociative extracts of foetal bovine nuchal ligament. Evidence for a relationship with type VI collagen

1984 ◽  
Vol 220 (2) ◽  
pp. 395-403 ◽  
Author(s):  
K R Knight ◽  
S Ayad ◽  
C A Shuttleworth ◽  
M E Grant
Keyword(s):  
Amino Acid ◽  
Ion Exchange ◽  
Amino Acid Analysis ◽  
Gel Filtration ◽  
Enzyme Digestion ◽  
Type Vi Collagen ◽  
Pepsin Digestion ◽  
Bacterial Collagenase ◽  
Collagenase Digestion ◽  
Nuchal Ligament

A collagenous glycoprotein (Mr 140000) was isolated from dissociative extracts of foetal bovine nuchal ligament and purified by a combination of ion-exchange and gel-filtration chromatography. This glycoprotein (designated MFPI) exists as a large-Mr disulphide-bonded aggregate in the absence of a reducing agent. The purified glycoprotein was shown to contain about 6% (w/w) carbohydrate, mostly as galactose, glucose and mannose. Amino acid analysis showed the presence of hydroxyproline and hydroxylysine, indicative of its collagenous nature. The collagenous nature of this glycoprotein was further investigated by enzyme digestion. Pepsin digestion produced three major fragments, which were identical with peptides of type VI collagen. Bacterial-collagenase digestion of the unreduced glycoprotein also produced several discrete peptides. However, reduction of the glycoprotein before bacterial-collagenase digestion resulted in the degradation of these discrete peptides. Glycoprotein MFPI extracted in dissociative conditions appears to be a larger-Mr form of type VI collagen, believed to originate from microfibrillar components in the intact tissue.

Gal3p and Gal1p interact with the transcriptional repressor Gal80p to form a complex of 1:1 stoichiometry

2002 ◽  
Vol 363 (3) ◽  
pp. 515-520 ◽  
Author(s):  
David J. TIMSON ◽  
Helen C. ROSS ◽  
Richard J. REECE
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acid ◽  
Gel Filtration ◽  
Transcriptional Repressor ◽  
Galactose Metabolism ◽  
Genetic Switch ◽  
Sequence Identity ◽  
Genes Encoding ◽  
Small Molecule Ligands ◽  
High Degree

The genes encoding the enzymes required for galactose metabolism in Saccharomyces cerevisiae are controlled at the level of transcription by a genetic switch consisting of three proteins: a transcriptional activator, Gal4p; a transcriptional repressor, Gal80p; and a ligand sensor, Gal3p. The switch is turned on in the presence of two small molecule ligands, galactose and ATP. Gal3p shows a high degree of sequence identity with Gal1p, the yeast galactokinase. We have mapped the interaction between Gal80p and Gal3p, which only occurs in the presence of both ligands, using protease protection experiments and have shown that this involves amino acid residue 331 of Gal80p. Gel-filtration experiments indicate that Gal3p, or the galactokinase Gal1p, interact directly with Gal80p to form a complex with 1:1 stoichiometry.

scholarly journals The primary structure of a short neurotoxin homologue from Dendroaspis polylepis polylepis (Black mamba) venom

1984 ◽  
Vol 3 (3) ◽  
pp. 169-174
Author(s):  
F. J. Joubert
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Ion Exchange ◽  
Amino Acid Sequence ◽  
Primary Structure ◽  
Gel Filtration ◽  
Cysteine Residues ◽  
Complete Amino Acid Sequence

Toxin CM-7 was purified from black mamba venom by gel filtration on Sephadex G-50 followed by ion exchange chromotography on CM-cellulose. It contains 60 amino acids, including eight half-cystines. The complete amino acid sequence of toxin CM-7 has been elucidated. In toxin CM-7 the eleven structurally invariant amino acids of the neurotoxins and cardiotoxins are conserved, but it has only one of the five functionally invariant amino acids of the neurotoxins. The eight cysteine residues of toxin CM-7 are in the same locations as those in short neurotoxins of known structures and are presumed to be similarly cross-linked. The sequence of CM-7 is structurally homologous with the short neurotoxins, but it is less toxic.

A novel bacteriocin-like substance (BLIS) from a pathogenic strain of Vibrio harveyi

Microbiology ◽  
2005 ◽  
Vol 151 (9) ◽  
pp. 3051-3058 ◽  
Author(s):  
Sathish Prasad ◽  
Peter C. Morris ◽  
Rasmus Hansen ◽  
Philip G. Meaden ◽  
Brian Austin
Keyword(s):  
Amino Acid ◽  
Vibrio Fischeri ◽  
Vibrio Harveyi ◽  
Gel Filtration ◽  
Anion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Proteinase K ◽  
Broth Culture ◽  
Two Dimensional Gel Electrophoresis ◽  
Flight Mass Spectrometry

Inter-strain and inter-species inhibition mediated by a bacteriocin-like inhibitory substance (BLIS) from a pathogenic Vibrio harveyi strain VIB 571 was demonstrated against four isolates of the same species, and one culture each of a Vibrio sp., Vibrio fischeri, Vibrio gazogenes and Vibrio parahaemolyticus. The crude BLIS, which was obtained by ammonium-sulphate precipitation of the cell-free supernatant of a 72 h broth culture of strain VIB 571, was inactivated by lipase, proteinase K, pepsin, trypsin, pronase E, SDS and incubation at ≥60 °C for 10 min. The activity was stable between pH 2–11 for at least 5 h. Anion-exchange chromatography, gel filtration, SDS-PAGE and two-dimensional gel electrophoresis revealed the presence of a single major peak, comprising a protein with a pI of ∼5·4 and a molecular mass of ∼32 kDa. The N-terminal amino acid sequence of the protein comprised Asp-Glu-Tyr-Ile-Ser-X-Asn-Lys-X-Ser-Ser-Ala-Asp-Ile (with X representing cysteine or modified amino acid residues). A similarity search based on the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) generated peptide masses and the N-terminal sequence did not yield any significant matches.

scholarly journals Heterologous Expression and Characterization of Two 1-Hydroxy-2-Naphthoic Acid Dioxygenases from Arthrobacter phenanthrenivorans

2011 ◽  
Vol 78 (3) ◽  
pp. 621-627 ◽  
Author(s):  
Elpiniki Vandera ◽  
Konstantinos Kavakiotis ◽  
Aristeidis Kallimanis ◽  
Nikos C. Kyrpides ◽  
Constantin Drainas ◽  
...  
Keyword(s):  
Amino Acid ◽  
Hydrophobic Interactions ◽  
Amino Acid Level ◽  
Gel Filtration ◽  
Amino Acid Sequences ◽  
Recombinant Enzymes ◽  
Content Type ◽  
Flight Mass Spectrometry ◽  
Naphthoic Acid ◽  
Dioxygenase Activity

ABSTRACTA protein fraction exhibiting 1-hydroxy-2-naphthoic acid (1-H2NA) dioxygenase activity was purified via ion exchange, hydrophobic interactions, and gel filtration chromatography fromArthrobacter phenanthrenivoranssp. nov. strain Sphe3 isolated from a Greek creosote-oil-polluted site. Matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) and tandem MS (MS-MS) analysis revealed that the amino acid sequences of oligopeptides of the major 45-kDa protein species, as analyzed by SDS-PAGE and silver staining, comprising 29% of the whole sequence, exhibited strong homology with 1-H2NA dioxygenase ofNocardioidessp. strain KP7. A BLAST search of the recently sequenced Sphe3 genome revealed two putative open reading frames, nameddiox1anddiox2, showing 90% nucleotide identity to each other and 85% identity at the amino acid level with theNocardiasp. homologue.diox1was found on an indigenous Sphe3 plasmid, whereasdiox2was located on the chromosome. Both genes were induced by the presence of phenanthrene used as a sole carbon and energy source, and as expected, both were subject to carbon catabolite repression. The relative RNA transcription level of the chromosomal (diox2) gene was significantly higher than that of its plasmid (diox1) homologue. Bothdiox1anddiox2putative genes were PCR amplified, cloned, and overexpressed inEscherichia coli. RecombinantE. colicells expressed 1-H2NA dioxygenase activity. Recombinant enzymes exhibited Michaelis-Menten kinetics with an apparentKmof 35 μM for Diox1 and 29 μM for Diox2, whereas they showed similar kinetic turnover characteristics withKcat/Kmvalues of 11 × 106M−1s−1and 12 × 106M−1s−1, respectively. Occurrence of twodiox1anddiox2homologues in the Sphe3 genome implies that a replicative transposition event has contributed to the evolution of 1-H2NA dioxygenase inA. phenanthrenivorans.

A NEUROPHYSIN FROM COD (GADUS MORRHUA) PITUITARY GLANDS: ISOLATION AND PROPERTIES

1968 ◽  
Vol 42 (1) ◽  
pp. 143-NP ◽  
Author(s):  
B. T. PICKERING
Keyword(s):  
Amino Acid ◽  
Ion Exchange ◽  
Amino Acid Composition ◽  
Arginine Vasopressin ◽  
Binding Capacity ◽  
Gel Filtration ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Maximum Binding Capacity ◽  
Pituitary Glands

SUMMARY A protein capable of binding neurohypophysial hormones has been isolated from cod pituitary glands using gel filtration and ion-exchange chromatography. The cod protein which was acidic and rich in cystine had an amino acid composition closely related to those of the mammalian neurophysins. It had a maximum binding capacity of 2·2 μmole/14mg. for oxytocin, 2·1 μmole/14 mg. for [8-arginine]-oxytocin and 1·1 μmole/14 mg. for [8-arginine]-vasopressin. Thus the cod protein had a greater capacity for the endogenous pressor-antidiuretic peptide than for the analogous mammalian hormone.

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