scholarly journals Anomalous behaviour of yeast isocitrate dehydrogenase during isoelectric focusing

1972 ◽  
Vol 129 (5) ◽  
pp. 1125-1130 ◽  
Author(s):  
John A. Illingworth
Keyword(s):  
Isoelectric Focusing ◽  
Isoelectric Point ◽  
Isocitrate Dehydrogenase ◽  
Band Pattern ◽  
Anomalous Behaviour ◽  
Multiple Forms ◽  
Isoelectric Points ◽  
The Mean ◽  
Crude Material

Isoelectric focusing of yeast isocitrate dehydrogenase apparently reveals a number of ‘isoenzymes’. These have isoelectric points near pH5.5 in crude material, but during purification the mean isoelectric point progressively rises to pH7.0 and the band pattern changes. The shift in isoelectric point during purification is apparently genuine, since it is also manifested in the electrophoretic and chromatographic properties of the enzyme. The multiple forms, however, are an artifact, generated by exposure of the enzyme to Ampholine, since their activities vary with the protein/Ampholine ratio and they cannot be observed in any system from which Ampholine is excluded. There are no detectable isoenzymes of yeast isocitrate dehydrogenase.

scholarly journals Isoelectric points of erabutoxins and monoacyl derivatives of erabutoxin b. Estimation of the pK values of amino groups in erabutoxins by using isoelectric-focusing data

1982 ◽  
Vol 203 (2) ◽  
pp. 427-433 ◽  
Author(s):  
N UI ◽  
C Takasaki ◽  
N Tamiya
Keyword(s):  
Isoelectric Focusing ◽  
Isoelectric Point ◽  
Amino Group ◽  
Amino Groups ◽  
Sea Snake ◽  
Isoelectric Points ◽  
Point Data ◽  
Laticauda Semifasciata ◽  
Erabutoxin B ◽  
Derivatives Of

The isoelectric points of erabutoxins a, b and c, neurotoxic proteins of a sea snake, Laticauda semifasciata, were determined by density-gradient isoelectric focusing. The same measurement was also made with monoacyl derivatives of erabutoxin b, in which each one of all amino groups had been either acetylated or propionylated. Erabutoxins a and b showed the same isoelectric point at pH 9.68. The values for [1-N alpha-acetyl-arginine]-, [15-N6-acetyl-lysine]-, [27-N6-acetyl-lysine]-, [47-N6-propionyl-lysine]- and [51-N6-acetyl-lysine]-erabutoxin b were at pH 9.52, 9.31, 9.45, 9.22 and 9.09 respectively, being definitely different from each other and lower than the value for the unmodified molecule. The isoelectric point of erabutoxin c, which is [51-asparagine]-erabutoxin b, was the same as that of [51-N6-acetyl-lysine]erabutoxin b. Assuming that no change in pK occurs on monoacylation, the pK values of amino groups in erabutoxin b were calculated from the isoelectric-point data. It is indicated that the pK values of zeta-amino groups differ markedly from each other and that the value of alpha-amino group is anomalously high.

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.

Isoelectric focusing of the human TSH receptor A subunit

1986 ◽  
Vol 6 (7) ◽  
pp. 685-689 ◽  
Author(s):  
F. A. Hashim ◽  
E. Davies Jones ◽  
R. D. Howells ◽  
B. Rees Smith
Keyword(s):  
Isoelectric Focusing ◽  
Isoelectric Point ◽  
Water Soluble ◽  
Tsh Receptor ◽  
Isoelectric Points ◽  
A Subunit ◽  
Receptor Antibodies ◽  
Tsh Receptor Antibodies ◽  
Charge Interactions

The water soluble A subunit of the human TSH receptor has been shown to have an isoelectric point of 5. As both TSH and TSH receptor antibodies have isoelectric points in the region of 8–10, charge-charge interactions must be of major importance in the binding of hormone or antibody to the TSH receptor A subunit.

Differentiation of Moniezia expansa and Moniezia benedeni (Eucestoda: Cyclophyllidea) by isoelectric focusing

1989 ◽  
Vol 67 (6) ◽  
pp. 1471-1475 ◽  
Author(s):  
M. R. Johnson ◽  
E. P. Hoberg
Keyword(s):  
Isoelectric Focusing ◽  
Intestinal Tissue ◽  
Banding Patterns ◽  
Isoelectric Points ◽  
The Mean ◽  
Moniezia Expansa ◽  
Host Tissues

Isoelectric focusing was performed on extracts from whole specimens of Moniezia expansa recovered from sheep and Moniezia benedeni recovered from cattle. The isoelectric focusing banding patterns for mature, postmature, early gravid, and gravid proglottids from individual cestodes were compared along with those for intestinal tissue from the respective hosts. Unique, reproducible banding patterns were characteristic of each species. In addition, the patterns for these cestodes were distinctly different from those for either of the host tissues. The tracks of the gels were scanned and values for isoelectric points were assigned for the dominant peaks characteristic of each species. A range based on the mean ± 2 SD was then assigned to each of these peaks for future comparisons.

scholarly journals Multiple forms of γ-butyrobetaine hydroxylase (EC 1.14.11.1)

1984 ◽  
Vol 223 (1) ◽  
pp. 119-127 ◽  
Author(s):  
S Lindstedt ◽  
I Nordin
Keyword(s):  
Isoelectric Focusing ◽  
Sodium Dodecyl Sulphate ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Human Kidney ◽  
Polyacrylamide Gels ◽  
Multiple Forms ◽  
Bacterial Enzyme ◽  
Isoelectric Points ◽  
Liver And Kidney

gamma-Butyrobetaine hydroxylase [4-trimethylaminobutyrate, 2-oxoglutarate:oxygen oxidoreductase (3-hydroxylating), EC 1.14.11.1] from human kidney was resolved into three forms by chromatofocusing. After further chromatography on an anion-exchanger, each form appeared as a single band on electrophoresis in polyacrylamide gel containing sodium dodecyl sulphate. The isoelectric points of isoenzymes 1, 2 and 3 were 5.6, 5.7 and 5.8 respectively, as estimated by isoelectric focusing. Their specific activities were 17-29 mu kat/g of protein. The concentrations of the three isoenzymes were about equal, possibly slightly lower for isoenzyme 1. The requirement for Fe2+ and the Km values for gamma-butyrobetaine and 2-oxoglutarate were about the same for the different enzyme forms. L- and D-Carnitine caused decarboxylation of 2-oxoglutarate to the same extent (8 and 29%) with the three forms. The enzyme forms had the same mass, 64 kDa, as determined by gel filtration in nondenaturing media. The same subunit mass, 42 kDa, was obtained for the multiple forms by electrophoresis in polyacrylamide gels containing sodium dodecyl sulphate. Isoenzyme 2 was resolved into two protein bands by isoelectric focusing in polyacrylamide gels containing urea. Isoenzyme 1 contained only one of these bands and isoenzyme 3 the other. The three enzyme forms of gamma-butyrobetaine hydroxylase thus appear to be dimeric combinations of two subunits differing in charge but not in size. gamma-Butyrobetaine hydroxylase from crude extracts of human, rat and calf liver was also separated into multiple forms by a chromatofocusing technique. The isoenzyme pattern was the same in human liver and kidney. The technique used to resolve the mammalian enzymes gave no evidence for the presence of multiple forms of the bacterial enzyme from Pseudomonas sp. AK 1.

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.

scholarly journals Isoelectric focusing of glutathione S-transferases from rat liver and kidney

1978 ◽  
Vol 175 (3) ◽  
pp. 937-943 ◽  
Author(s):  
Barbara F. Hales ◽  
Valerie Jaeger ◽  
Allen H. Neims
Keyword(s):  
Substrate Specificity ◽  
Isoelectric Focusing ◽  
Transferase Activity ◽  
Sprague Dawley ◽  
Substrate Specificities ◽  
Liver Cytosol ◽  
Sprague Dawley Rats ◽  
Isoelectric Points ◽  
Liver And Kidney ◽  
Glutathione S Transferases

The glutathione S-transferases that were purified to homogeneity from liver cytosol have overlapping but distinct substrate specificities and different isoelectric points. This report explores the possibility of using preparative electrofocusing to compare the composition of the transferases in liver and kidney cytosol. Hepatic cytosol from adult male Sprague–Dawley rats was resolved by isoelectric focusing on Sephadex columns into five peaks of transferase activity, each with characteristic substrate specificity. The first four peaks of transferase activity (in order of decreasing basicity) are identified as transferases AA, B, A and C respectively, on the basis of substrate specificity, but the fifth peak (pI6.6) does not correspond to a previously described transferase. Isoelectric focusing of renal cytosol resolves only three major peaks of transferase activity, each with narrow substrate specificity. In the kidney, peak 1 (pI9.0) has most of the activity toward 1-chloro-2,4-dinitrobenzene, peak 2 (pI8.5) toward p-nitrobenzyl chloride, and peak 3 (pI7.0) toward trans-4-phenylbut-3-en-2-one. Renal transferase peak 1 (pI9.0) appears to correspond to transferase B on the basis of pI, substrate specificity and antigenicity. Kidney transferase peaks 2 (pI8.5) and 3 (pI7.0) do not correspond to previously described glutathione S-transferases, although kidney transferase peak 3 is similar to the transferase peak 5 from focused hepatic cytosol. Transferases A and C were not found in kidney cytosol, and transferase AA was detected in only one out of six replicates. Thus it is important to recognize the contribution of individual transferases to total transferase activity in that each transferase may be regulated independently.

Morph-Specific Proteins in Pollen and Styles of Distylous Turnera (Turneraceae)

Genetics ◽  
1997 ◽  
Vol 146 (2) ◽  
pp. 669-679
Author(s):  
Andreas Athanasiou ◽  
Joel S Shore
Keyword(s):  
Isoelectric Focusing ◽  
Isozyme Markers ◽  
Unlinked Locus ◽  
Full Expression ◽  
Isoelectric Points ◽  
Isozyme Loci ◽  
Specific Proteins ◽  
S Allele

We used nondenaturing isoelectric focusing (IEF) in a survey of plants from 11 populations to identify style and pollen proteins unique to the short-styled morph of Turnera scabra, T. subulata and T. krapovickasii. Three protein bands [approximately isoelectric points (pIs) 6.1, 6.3 and 6.5] were found only in styles and stigmas of short-styled plants while two bands (approximately pIs 6.7 and 6.8, M  r 56 and 59 kD) occur only in pollen of short-styled plants. Some of these bands appear very late in development, within 24 hr before flowering. Two isozyme loci were mapped to an 8.7 cM region spanning the distyly locus. Using these isozyme markers we identified progeny exhibiting recombination adjacent to the distyly locus. No recombinants between the distyly locus and the locus or loci controlling the presence of the short-styled morph-specific proteins were obtained. This suggests that the loci encoding these proteins are either extremely tightly linked to the distyly locus and in complete disequilibrium with the S allele or exhibit morph-limited expression. Crosses to a plant showing an unusual style protein phenotype demonstrated that an additional unlinked locus is required for full expression of the style proteins. The function of the morph-specific proteins is unknown

Sign in / Sign up

Export Citation Format

Share Document