Isoelectric focusing of insulin, 125I-insulin and the 125I-insulin-antibody complex

1979 ◽  
Vol 44 (6) ◽  
pp. 1828-1834
Author(s):  
Asja Šiševa ◽  
Jiřina Slaninová ◽  
Tomislav Barth ◽  
Stephan P. Ditzov ◽  
Luben M. Sirakov
Keyword(s):  
Isoelectric Focusing ◽  
Polyacrylamide Gel ◽  
The Other ◽  
Insulin Antibody ◽  
Isoelectric Points ◽  
Antibody Complex ◽  
And Storage ◽  
Acid Region ◽  
Three Components

Isoelectric focusing on polyacrylamide gel columns of three native crystalline commercial preparations of insulin and 125I-labelled insulin was carried out. All the compounds studied contained three components of different isoelectric points. The largest fraction, having pI 5.60 ± 0.05, was common to all preparations. The other two fractions were situated in the acid region of pH between pI 4.5 and 5.2. The presence of these fractions is explained by the contamination of crystalline insulins by proinsulin and by the formation of des-amido derivatives during the dissolving and storage of insulin samples, and, in case of labelled insulin, also by the presence of heavily iodinated insulin and contaminating components. The isoelectric focusing of the complex 125I-insulin-antibody showed a peak of radioactivity having pI 6.15 ± 0.05.

Resolution of rat renin substrates by isoelectric focusing

1977 ◽  
Vol 55 (8) ◽  
pp. 869-875 ◽  
Author(s):  
A. A. Faiers ◽  
A. Y. Loh ◽  
D. H. Osmond
Keyword(s):  
Isoelectric Focusing ◽  
Substrate Concentration ◽  
Rat Plasma ◽  
The Other ◽  
Multiple Forms ◽  
High Substrate Concentration ◽  
Broad Distribution ◽  
Renin Substrate ◽  
Isoelectric Points ◽  
Different Sources

Pooled plasmas from normal or binephrectomized rats and perfusates of isolated livers were used as sources of renin substrate for isoelectric focusing. After desalting, preliminary fractionation (plasma only), and concentration, the preparations were focused in a pH 3–10 gradient on 20-cm glass plates layered with Sephadex slurry. The pH 4–6 region, containing all the substrate, was scraped from this plate and refocused in a pH 4–6 gradient. Substrate content of 1-cm strips of slurry from half of the plate was determined by both radioimmunoassay and bioassay of angiotensin resulting from incubation with added renin. Corresponding strips from the other half of the plate were incubated without renin as a control for any preformed angiotensin. The asymmetry and broad distribution (pH 4–5) of substrate from different sources suggested the existence of more than one form. Higher resolution achieved by using the high substrate concentration of postnephrectomy plasma and 0.5-cm strips of slurry on 20-cm or 40-cm plates revealed peaks and shoulders of substrate activity. Our data suggest that multiple forms of substrate are synthesized by the liver and circulate in plasma. Postnephrectomy rat plasma appears to contain relatively more substrate(s) with higher isoelectric points than in normal plasma, possibly an accumulation of forms ordinarily degraded by endogenous renal renin.

scholarly journals Formation and properties of flavoprotein-cytochrome hybrids by recombination of subunits from different species

1985 ◽  
Vol 231 (2) ◽  
pp. 383-387 ◽  
Author(s):  
S C Koerber ◽  
D J Hopper ◽  
W S McIntire ◽  
T P Singer
Keyword(s):  
Catalytic Activity ◽  
Steady State ◽  
Isoelectric Focusing ◽  
Dissociation Constants ◽  
The Other ◽  
High Dilution ◽  
Steady State Kinetic ◽  
Isoelectric Points ◽  
State Kinetic ◽  
Enzyme Species

p-Cresol methylhydroxylases from four different pseudomonads differ in their isoelectric points and, to a lesser extent, in Mr values and substrate specificity. The enzymes from three species were isolated in homogeneous form, then resolved into their flavoprotein and cytochrome subunits, and the subunits were recombined to yield the nine possible hybrids (i.e. three intraspecies and six interspecies). The resulting flavocytochromes showed extensive similarities in steady-state kinetic parameters and in the dissociation constants of their subunits. Evidence is also presented that a fourth type of p-cresol methylhydroxylase, from Pseudomonas putida (N.C.I.B. 9869, form ‘B’), the subunits of which cannot be isolated by the isoelectric focusing technique used to separate the subunits of the other flavocytochromes, nevertheless dissociates slowly at high dilution. The dissociation is reflected by a decline of catalytic activity with time. This process for the ‘B’ enzyme is prevented by the presence of substrate or an excess of a cytochrome subunit isolated from another enzyme species. Incubation of the dissociated subunits with p-cresol brings about extensive, albeit incomplete, re-association and regeneration of activity.

Ontogeny of Hamster Haemoglobins Determined by Isoelectric Focusing in Polyacrylamide Gel

1974 ◽  
Vol 16 (3) ◽  
pp. 677-686
Author(s):  
THALIA BOUSSIOS ◽  
J. F. BERTLES
Keyword(s):  
Isoelectric Focusing ◽  
Gel Electrophoresis ◽  
Laboratory Animal ◽  
Polyacrylamide Gel ◽  
The Other ◽  
Suitable Model ◽  
Small Laboratory Animal ◽  
Small Laboratory ◽  
Foetal Haemoglobin ◽  
Haemoglobin Synthesis

Haemoglobin ontogeny in an inbred strain of the golden hamster was determined from 12 days in gestation to adulthood. Haemoglobins, separated by isoelectric focusing in polyacrylamide gel in a linear pH gradient (7.0 to 8.0), were quantified by optical density scanning (420 nm) of the unstained gels. Three species (adult haemoglobins) increase in proportion, one (15 % at 12 days gestation) becomes dominant (85 % in adults), and two (nearly absent at 12 days gestation) increase and exist as minor species in adults. Two species (foetal haemoglobins) decrease rapidly, one (37 % at 12 days gestation) to trace levels, the other (24 % at 12 days gestation) to a persistent 3 % in adults. Isoelectric-focused haemoglobins were eluted individually and re-identified by isoelectric focusing, disk-gel electrophoresis, and vertical gel electrophoresis. Gel exclusion studies ruled out the possibility that any one haemoglobin species is a polymer of another. Haemoglobin proportions determined by the technique used here, isoelectric focusing, are very reproducible, both from sample to sample and from hamster to hamster of any specific age. The presence of a foetal haemoglobin fraction persisting at significant levels into adulthood suggests that this small laboratory animal is a suitable model for studies on foetal haemoglobin synthesis.

Fractionation and further characterization of granulocytic and monocytic alpha-naphthyl acetate (ANAE) esterases.

1984 ◽  
Vol 32 (6) ◽  
pp. 579-584 ◽  
Author(s):  
C S Scott ◽  
D Hough ◽  
A G Bynoe ◽  
D B Jones ◽  
B E Roberts
Keyword(s):  
Cell Differentiation ◽  
Isoelectric Focusing ◽  
Myeloid Cell ◽  
The Other ◽  
Monocytic Differentiation ◽  
Isoelectric Points ◽  
Monocytic Lineage ◽  
Naphthyl Acetate ◽  
Nonspecific Esterases

Following characterization of myeloid nonspecific esterases by isoelectric focusing (IEF), two main groups of alpha-naphthyl acetate (ANAE) esterase isoenzymes were defined and fractionated from cytoplasmic extracts by chromato focusing techniques according to differences in their isoelectric points (pI). The first of these ANAE enzyme groups was common to leukocytes of both granulocytic and monocytic lineage, while the other, which characteristically comprised a group of isoenzymes within the pI range 5.5-6.1, was specifically associated with monocytic differentiation. The properties of the two purified ANAE enzyme fractions were compared by inhibition (heat and sodium fluoride) and further electrophoretic studies, and the results discussed in relation to the cytochemical characterization of these enzymes as markers of specific myeloid cell differentiation.

scholarly journals Characterization of the estrogen-induced uterine peroxidase by microelectrophoresis.

1978 ◽  
Vol 26 (5) ◽  
pp. 382-390 ◽  
Author(s):  
C C Phillips Burnett ◽  
W A Anderson ◽  
R Rüchel
Keyword(s):  
Molecular Weight ◽  
Isoelectric Focusing ◽  
Large Subunit ◽  
The Other ◽  
Two Dimensional ◽  
Neuraminidase Treatment ◽  
Gradient Electrophoresis ◽  
Isoelectric Points ◽  
Uterine Fluid

Estrogen-dependent peroxidase from rat uterine fluid has been investigated by microelectrophoretic techniques. The molecular weight of the enzyme has been determined in the range of 100,000 by using polyacrylamide gradient gels in the absence and presence of nonionic and anionic detergent. The isoelectric points are located between pH 4.5 and 5.9. Employing the two-dimensional combination of isoelectric focusing and gel gradient electrophoresis, the enzyme was separated into two subunits, one having a molecular weight of 70,000, the other less than 20,000. The large subunit has slight enzymatic activiy, while the smaller subunit may be responsible for the charge difference in the holoenzyme pattern. The glycoprotein pattern of the uterine fluid peroxidase is further defined by its separation by affinity chromatography using a concanavalin A-Sepharose column and by its susceptibility to neuraminidase treatment.

A METHOD FOR ISOLATION BY GEL ELECTROFOCUSING OF ISOHORMONES B AND C OF HUMAN PROLACTIN FROM AMNIOTIC FLUID

1980 ◽  
Vol 84 (1) ◽  
pp. 125-133 ◽  
Author(s):  
MENASHE BEN-DAVID ◽  
ANDREAS CHRAMBACH
Keyword(s):  
Amniotic Fluid ◽  
Isoelectric Focusing ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Polyacrylamide Gel ◽  
Constant Ph ◽  
Human Prolactin ◽  
Isoelectric Points ◽  
Carrier Ampholytes

Human prolactin from amniotic fluid, consisting of isohormones B and C (major), was radio-iodinated after storage of the hormone for 3 years at −70 °C, and yielded a Ferguson plot in polyacrylamide gel electrophoresis that was indistinguishable from the original except that the zones of isohormone B and C were fused. However, isohormones B and C of 125I-labelled human prolactin were separated on isoelectric focusing in polyacrylamide gel, using Ampholine carrier ampholytes (pI range 5–8), taurine (pI 5·1) as anolyte and β-alanine (pI 6·9) as catholyte. After 20 h of electrofocusing at 0–4 °C, 1000 V, both isohormones reached constant pH (isoelectric) positions on the gel. The apparent isoelectric points of human prolactin B and C were 5·96 and 5·62 Micro-preparative gel electrofocusing followed by excision and re-electrofocusing of the gel slices containing human prolactin B and C, yielded zones of homogeneous isohormones B and C.

Penicillocarboxypeptidase-S, a Nonspecific SH-Dependent Exopeptidase

1972 ◽  
Vol 50 (12) ◽  
pp. 1297-1310 ◽  
Author(s):  
S. R. Jones ◽  
T. Hofmann
Keyword(s):  
Isoelectric Focusing ◽  
Culture Medium ◽  
Iodoacetic Acid ◽  
Cysteine Residue ◽  
The Other ◽  
Ph Optimum ◽  
Penicillium Janthinellum ◽  
Diisopropyl Phosphorofluoridate ◽  
Isoelectric Points ◽  
Inhibition Studies

An extracellular carboxypeptidase with a pH optimum of between 4 and 5 has been isolated from the culture medium of Penicillium janthinellum. Like penicillopepsin, this enzyme is only released when the stationary phase of growth is reached. The enzyme has been purified by affinity chromatography on phenylbutylamine-Sepharose followed by isoelectric focusing. The enzyme was found to be present in two forms with isoelectric points of 3.70 and 3.77. It has a molecular weight of about 48 000 and is inhibited by 10−7 M p-hydroxymercury benzoic acid. It is not inhibited by the metal chelators EDTA, o-phenanthroline, and 8-hydroxyquinoline, or by diisopropyl phosphorofluoridate. The presence of a single cysteine residue has been demonstrated by labelling the enzyme with 14C-iodoacetic acid. The action of the enzyme on glucagon, the S-sulfo-B-chain of insulin, and on penicillopepsin at pH 4.7 shows that it is a nonspecific carboxypeptidase and releases C-terminal proline, lysine, and arginine, as well as the other amino acids. Glycine appears to be the slowest residue to be released. Carbobenzoxy-L-glutamyl-L-tyrosine, the substrate used for routine assays, is hydrolyzed very rapidly. The enzyme also slowly hydrolyzed Leu–Phe, and splits glycine from Leu–Gly–Gly.A second enzyme with carbobenzoxy-L-glutamyl-L-tyrosine activity is present in the growth medium. It has an isoelectric point of 4.72 on an isoelectric focusing column. Preliminary inhibition studies of a partially purified preparation suggest that it differs from the other enzyme.

EFEK PENAMBAHAN EKSTRAK AIR JAHE (Zingiber officinale Roscoe) DAN PENYIMPANAN DINGIN TERHADAP MUTU SENSORI IKAN TUNA (Thunnus albacores)

2010 ◽  
Vol 6 (1) ◽  
pp. 36
Author(s):  
Silvana Dinaintang Harikedua
Keyword(s):  
Sensory Quality ◽  
Zingiber Officinale ◽  
The Other ◽  
Ginger Extract ◽  
Other Hand ◽  
Storage Treatment ◽  
Zingiber Officinale Roscoe ◽  
Hedonic Scale ◽  
And Storage

The objective of this study was to investigate the effect of ginger extract addition and refrigerate storage on sensory quality of Tuna through panelist’s perception. Panelists (n=30) evaluated samples for overall appearance and flavor attribute using hedonic scale 1–7. The sample which is more acceptable by panelists on flavor attributes having 3% gingers extract and storage for 3 days. The less acceptable sample on flavor attribute having 0% ginger extract and storage for 9 days. On the other hand, the sample which is more acceptable by panelists on overall appearance having 0% ginger extract without storage treatment. The less acceptable sample on overall appearance having 3% ginger extract and storage for 9 days.

A System Architecture for Data Storage in the Cloud

2015 ◽  
Vol 3 (2) ◽  
Author(s):  
Maryam Hammami ◽  
Hatem Bellaaj
Keyword(s):  
Data Storage ◽  
System Architecture ◽  
Cloud Storage ◽  
The Other ◽  
Important Data ◽  
Work In Progress ◽  
Flexible System ◽  
Other Hand ◽  
Data Manager ◽  
And Storage

The Cloud storage is the most important issue today. This is due to a rapidly changing needs and a huge mass of varied and important data to back up. In this paper, we describe a work in progress and propose a flexible system architecture for data storage in the Cloud. This system is centered on the Data Manager module. This module provides various functions such as the dispersion of data in fragments, encryption and storage of fragments... etc. This architecture proves to be very relevant. It ensures consistency between different components. On the other hand, it ensures the security and availability of data.

Sign in / Sign up

Export Citation Format

Share Document