Transferrin Variants Research Articles

Electrophoretic transferrin variation in fur seals (Arctocephalus spp.) at Marion Island.

1. Variation in transferrin types were investigated in sympatric populations of two fur seal species which are undergoing limited hybridization at Marion Island. 2. Vertical polyacrylamide gel electrophoresis of serum was performed to demonstrate the transferrin types. 3. The two species appear fixed for alternative alleles: A. tropicalis being homozygous for the fast allele, and A. gazella (with one exception) being homozygous for the slow allele, indicating low gene flow between these two species. The single hybrid tested was homozygous for the slow allele. 4. The use of electrophoretically determined transferrin variation holds promise for the investigation of these and other sympatric fur seal populations.

Difference in content ratio of components among horse serum transferrin variants.

Each of five genetic variants of horse serum transferrin (Tf), D, F, H, O, and R, was separated into two bands by polyacrylamide gel isoelectric focusing (PAGIEF). The more acidic band, termed component a, was more abundant than the other one, termed component b, in all variants. Components a and b of TFO variant were immunologically indistinguishable from each other by double immunodiffusion test. Determination of the content ratio of component a to component b in each variant revealed that the variants were classified into two groups: one group (D, F, and H) had a relatively high ratio within a range from 3.4 to 4.0 and the other group (O and R) had a relatively low ratio of 1.8 to 2.3. From these results and reference data on carbohydrate compositions of components a and b, it was proposed that there is a difference in glycosylation between the two groups.

Transferrin and mitochondrial aspartate aminotransferase in young adult alcoholics

Two recently proposed biochemical markers of alcoholism, namely, quantitation of plasma transferrin variant (Tf 5.7) and the ratio of plasma mitochondrial aspartate aminotransferase (m-AspAT) to total AspAT (t-AspAT), were tested for their ability to detect young adult alcoholics. Another commonly used biochemical test, namely, activity of plasma gamma glutamyltransferase (GGT) was included as a comparison. Although mean values of GGT, TF 5.7, total transferrin (Tf tot), m-AspAT and t-AspAT in alcoholics were significantly higher than those in controls, there were too many overlapping values in each test between alcoholics and controls to render any of these tests suitable as a marker for young adult alcoholics. Depending on cut-off limits, the sensitivity of each test ranged from 0–52% and the specificity ranged from 80–97%. Moreover, the m-AspAT/ t-AspAT and Tf 5.7 Tf tot ratios were not significantly different between alcoholics and controls. A stepwise linear discriminant function analysis of all the variables resulted in a slight increase in classification sensitivity (66%) but a decrease in specificity (77%). The relatively short duration (mean = 5.6 years) of heavy alcohol intake and the time elapsed (mean = 5.8 days) since the alcoholics last consumed alcohol very likely contributed to the low sensitivity. Young adults might also be more resilient with regard to the damaging biochemical effects of ethanol. Abnormal biochemical values might reverse to normal values much more quickly in young adult alcoholics than in those who are older and have more years of alcohol abuse.

A new transferrin variant from Iran (Tf B-Iran): Review of 36 variant alleles

A total of 2581 serum samples collected from five population groups of Iran was studied for electrophoretic variations of the transferrin (Tf). Besides the common phenotype Tf C the authors could observe 41 individuals with rare Tf types: CB1, CB2, CD1, CDChi, CD2. In addition to these Tf types two individuals with a new Tf B variant were observed. This new variant was found in the Dezfooli sample and was designated as Tf B-Iran. The electrophoretic position of this variant is described, and all the hitherto known Tf variants are reviewed.

Characterization of the amino acid change in a transferrin variant

in the serum before addition of the iron or indium, however, after addition of the metal ions the transferrins migrated as a single fast-moving species. Urea/gel electrophoresis has also been used to investigate the ability of In( I l l ) to displace Zn(l l ) from ovotransferrin. Although zinc binds specifically to the protein, the zinc complex was unstable on the polyacrylamide gel and migrated as the metal-free protein. When a two-fold excess of indium chloride was added to zinc-saturated ovotransferrin the zinc was displaced completely as judged by the mobility of the protein upon urea/gcl clectrophoresk. T h e results of this study show that In(llI) forms tight complexes with both human serum transferrin and hen ovotransferrin which retain the indium upon electrophoresis in the presence o f 6 M-urea. This technique can therefore be used to investigate the ability of transferrin to remove indium from chelators whch are currently being developed for use as indium-labelled pharmaceuticals.

Synthesis of serum proteins by cultured aggregates from endodermal cells of the area opaca of the primitive streak chick embryo.

The pattern of serum protein synthesis and secretion in aggregates of extraembryonic endoderm cells (EEC) from the area opaca of primitive streak chick embryos was studied. EEC aggregates were cultured for various time intervals and serum proteins were detected using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and fluorography. Serum proteins were identified based on their comigration with reference proteins from 4 day chick embryo serum and with reference proteins from egg white albumen and chicken serum. A number of serum proteins were detected in EEC aggregates including: two variants of immunoglobulin (IgG), four variants of transferrin, a protein with a molecular weight of 66 500 which may correspond to α globulins, prealbumin, and a protein with a molecular weight of 38 600 (serum protein 11) which remains unidentified. These proteins were also detected in the culture medium. The banding profiles of EEC extracts and culture medium were compared over various time intervals of culture (6, 18 and 30 h). The IgGs, transforms and serum protein 11 decreased in concentration in EEC extracts over the culture interval. These proteins as well as prealbumin, were detected in the culture medium. A number of proteins were synthesized by EEC, as determined by radiolabelled amino acid incorporation. All of the labelled serum proteins were detected in the culture medium, not in EEC extracts. These results suggest that serum proteins are synthesized by EEC then rapidly released into the medium. Labelled serum proteins detected in the culture medium include prealbumin and an unidentified serum protein (serum protein 14) which migrates with the tracking dye, both synthesized early in culture (6 h), and transferrin which was synthesized later (18 h) during culture.

Electrophoretic variants of serum transferrin in wild pig populations of Japan.

Variants of serum transferrin in Japanese wild pig, Sus scrofa leucomystax, and Ryukyu wild pig, S.s. riukiuanus, of Japan were investigated by using starch gel electrophoresis. Five phenotypes, TfB, BC, C, CX and X, were observed, of which two, TfCX and X, are new variants. Comparison of gene frequency estimates which were calculated for each population showed remarkable geographic differences among several populations of these two subspecies.

Influence of food intake on the 24-hr variations of plasma iron concentration in the rabbit.

Circadian variations in plasma iron levels were first reported in humans in 1937. Influences of the sleeping pattern and of plasma cortisol and adrenaline levels on these variations as well as the reproducibility of the phenomenon itself are discussed controversially in the literature. The influence of food intake, however, was not considered in most of the studies and is therefore subject of this investigation. Circadian plasma iron and plasma transferrin variations were determined in rabbits and compared under free access to food and under starvation (caecotrophy was not prevented). Population-mean-cosinor analysis of circadian plasma iron concentrations showed similar variations in the fed and starved condition (mesor: 116.6 micrograms/dl vs 118.1 micrograms/dl, acrophase 0752 hr vs 0728) except for a significant increase of the circadian amplitude under free access to food (30.9 micrograms/dl vs 22.3 micrograms/dl, P less than 0.05). There was no variation in plasma transferrin, which shows that 24 hr variations in plasma iron are not caused by modulation of plasma transferrin. These findings demonstrate a circadian rhythm for plasma iron, the amplitude of which is increased by food intake.

Variation of Transferrin and Esterase in Sera of Dogs

Transferrin (Tf), arylesterase (ArE) and another esterase (Es) have been studied in sera from 1023 dogs by the use of isoelectric focusing (IEF) in polyacrylamide gels. Tf types were determined after protein staining in gels of pH range 5–6 and 5–7. The expression of Tf types as measured by strength of bands varied considerably. The Tf band patterns are explained by the occurrence of the 4 codominant alleles, TfF, TfM ΤfM2 and TfS of which TfM1 and TfM2are common. Some breeds had similar gene frequencies, others differed considerably. For determination of ArE types specifically stained gels of pH range 4.2–4.9 and 4.0–6.5 were employed. The ArE phenotypes appeared as multiple band patterns of which the individual bands varied considerably in strength. Atypical ArE patterns were observed in dogs suffering from certain diseases. The normal ArE phenotypes are explained by a total of 7 codominant alleles of which ArEN and ArET have not been previously described. Gene frequencies varied between breeds. For the other esterase (Es) the appearance and position of bands indicate at least 2 alleles in this system.

Progressive Desialidation of Human Transferrin

Transferrin is a serum glycoprotein which contains four sialic acid residues located at the end of two branched carbohydrate structures. The presence of these four acidic residues influences the electrophoretic mobility of the transferrin molecule. Alterations in the electrophoretic mobility of transferrin may be encountered in forensic science case work, particularly in association with postmortem samples. These altered transferrins usually appear in a highly stylized "ladder" banding pattern. To determine whether these altered transferrins are the result of sialic acid removal, serum samples of known transferrin type were treated with neuraminidase. These experiments support the hypothesis that the "ladder" banding pattern of transferrin observed in some case samples is due to the removal of sialic acid residues by bacterial or endogenous neuraminidase. These studies also demonstrate that partially desialidated transferrin variants cannot be clearly typed until the sialic acid is completely stripped from the transferrin molecule. Reliable typing of partially desialidated samples can be accomplished by treating these samples with neuraminidase.

Transferrin variation and genetic structure of reindeer populations in Scandinavia

<p>Polyacrylamide gel electrophoresis was used to analyse transferrin variation in herds of semi-domestic reindeer from Scandinavia. The results are compared with previously reported values for other populations of both semi-domestic and wild reindeer using the same techniques as in the present study. In all populations the number of alleles was high, ranging from seven to eleven, and the heterozygosity was correspondingly high, with a mean of 0.749. This high genetic variation in all populations suggests that inbreeding is not widespread among Scandinavian reindeer. The pattern of allele frequency distribution indicates a high degree of genetic heterogeneity in the transferrin locus, both between the different semi-domestic herds and between the different wild populations. The mean value of genetic distance was 0.069 between semi-domestic herds and 0.091 between wild populations. Between semi-domestic and wild populations the genetic distance was particularly high, with a mean of 0.188. This high value was mainly due to a different pattern in the distribution of the two most common transferrin alleles: Tfu was most common among semi-domestic herds, while TfEI was most common among wild populations. These differences in transferrin allele distribution are discussed in relation to possible different origins of semi-domestic and wild reindeer in Scandinavia, or alternatively, to different selection forces acting on transferrin genotypes in semi-domestic and wild populations.</p><p>Transferrin-variasjon og genetisk struktur hos rein i Skandinavia.</p><p>Abstact in Norwegian / Sammendrag: Transferrin-variasjon i tamreinflokker ble analysert ved hjelp av polyacrylamid gel elektroforese. Resultatene er sammenlignet med verdier som tidligere er beskrevet for både tamrein og villrein hvor det ble benyttet samme metode som i denne undersøkelsen. I alle populasjonene ble det registrert et høyt antall alleler (7-11) og heterozygositeten var tilsvarende høy med en middelverdi på 0.749. Denne høye graden av genetisk variasjon i alle undersøkte populasjoner indikerer at det ikke foregår mye innavl blant rein i Skandinavia. Utbredelsen av de enkelte allelene viste høy grad av genetisk oppdeling i transferrin-locuset mellom flokker av både tamrein og villrein. Middelverdien for genetisk avstand var 0.069 mellom tamreinflokker og 0.091 mellom villreinflokker. Særlig stor genetisk avstand (middelverdi 0.188) ble funnet mellom tamrein og villrein. Denne store forskjellen skyldes i stor grad forskjellig mønster i utbredelsen av de to vanligste allelene: Tf' var mest vanlig blant tamrein og Tf1' var mest vanlig blant villrein. Denne forskjellen er diskutert i relasjon til forskjellig opprinnelse av tamrein og villrein og alternativt, i relasjon til forskjellig seleksjonskrefter som virker på transferrin genotyper i tamrein og villrein.</p>

Neue Methoden zur Diagnostik von Liquorfisteln mit Hilfe von β2-Transferrin oder Präalbumin - Grundlagen und Methodik

Two new methods were developed to identify pure CSF and CSF in nasal secretion. One method is based on measurement of beta 2-transferrin, a transferrin variant found only in CSF, whereas the other one is based on determination of the albumin-prealbumin ratio. The determination of beta 2-transferrin is made in two steps. Firstly, the protein variants beta 1- and beta 2-transferrin are isolated selectively from the complex protein mixture by an affinity procedure. The transferrin variants are then separated by a highly resolving polyacrylamide gelelectrophoresis and identified by staining. Advantages of the method are that blood or nasal secretion does not disturb the test and that beta 2-transferrin can be identified with high accuracy. Therefore, CSF can be differentiated unambiguously from other secretions. The determination of the ratios of concentration of the proteins albumin and prealbumin by immunoelectrophoresis is a further possibility to identify pure or almost pure CSF. This sensitive method, which is fast and easy to perform, is very helpful in corroborating the diagnosis of a CSF fistula. On the basis of this method one cannot get a false positive result, but a false negative one by high contamination of CSF with blood or nasal secretion. Therefore, the method of beta 2-transferrin determination must be carried out if there is any doubt.

Transferrin variants in Japan and New Zealand. Report of an unusually sialyzed TF variant.

The genetic polymorphism of transferrin (TF) was studied in 2,167 Japanese individuals and in 448 New Zealanders. The three TF C subtypes were identified, but TF C3 was absent from Japanese populations. In addition, three TF B and six TF D variants were observed, each of which occurred either in Japanese or in New Zealanders. Stepwise removal of N-acetyl-neuraminic acid (NANA) with neuraminidase revealed that the most cathodal variant TF DShinnanyo found in a Japanese family was characterized by having only two NANA residues.

Improved separation of the common transferrin variants in gels containing pH 5–7 Ampholines and HEPES

AbstractAn ultrathin‐layer polyacrylamide gel isoelectric focusing technique is described for obtaining highly resolved transferrin (Tf) subtypes. N‐2‐hydroxyethylpiperazine‐N′‐2‐ethanesulfonic acid (HEPES) buffer was used to flatten the pH gradient in gels containing pH 5–7 Ampholines. The result was increased separation distances for the TfC subtype bands. The method is effective even with the most recent lots of pH 5–7 Ampholines which were not capable of providing an appropriate environment for effective TfC subtyping.

A new transferrin allele in sheep

An electrophoretic variant of sheep transferrin, TfL, has been described. Transferrin L has been shown to be controlled by a single codominant allele, TfL, at the Tf locus. Transferrin L is electrophoretically distinguishable from the very similar transferrin TfKCzech. The value of gradient polyacrylamide gel electrophoresis for transferrin phenotyping in sheep is discussed.

The reliability of the use of para toluene sulfonic acid for simultaneous hydrolysis and quantitation of both N-acetyl-glucosamine and amino acids in human transferrins

Human transferrin preparations [l-3] isolated by preparative isoelectric focusing from normal and pathological sera show microheterogeneity which is caused by differences in the carbohydrate chain. Since transferrin variants are found in sera from alcoholics [4], patients with liver diseases [24] and patients with demyelinating disease [3], we wished to develop a method by which in one run, hydrolysis and quantitation, the amino acid and hexosamine composition can be determined and the transferrin variants can be compared. Recently, we described a method to determine neutral carbohydrates of transfertin on an adapted amino acid analyzer [5]. The transferrin subfractions did not only differ in sialic acid but also in galactose and mannose, and this makes it desirable to determine also the remaining sugar, N-acetyl-glucosamine. Since 1978, a number of papers [a-14] have been published describing the use of a normal amino acid analyzer with, in some cases, the simultaneous analysis of both hexosamines and amino acids [6-91. However problems arise with hydrolysis of both. One is aware of the fact that hexosamines were more easily destroyed in 6 mol/l HCl than most of the amino acids. In order to achieve a simple hydrolysis step followed by amino acid and hexosamine analysis, we have combined the hydrolysis method of Liu and Chang [15] who used 3 mol/l paratoluenesulfonic acid monohydrate (pTSA) at 1lO“C for 22, 48 and 72 h for the hydrolysis of amino acids and the method of Liu [16] who used 2 mol/l pTSA at 100°C for 2, 6, 12 and 24 h for the release of mannosamine phosphate from A polysaccharide. We now describe the amino acid and glucos-

Transferrin variation and evolution of Alaskan reindeer and caribou, Rangifer tarandus L.

Polyacrylamide gel electrophoresis was used to analyse transferrin variation in wild caribou (Rangifer tarandus granti) and domestic reindeer (R.t. tarandus) from Alaska. Eighteen alleles were detected in caribou and ten alleles were detected in reindeer. The most common allele was Tf 1 with a frequency of 0.304 and 0.408 in caribou and reindeer, respectively. The allele frequency distributions were significantly different in reindeer and caribou. This finding, together with the absence in reindeer of nine alleles present in caribou, suggests that little genetic exchange has taken place between caribou and reindeer in Alaska. The allele frequency distribution in Alaska caribou and reindeer are compared with those for other populations of caribou and reindeer. This comparison indicates that Alaskan caribou as well as Eurasian reindeer have evolved from a common ancesteral population different from the ancesteral population of Peary cairbou (R.t. pearyi) and Svalbard reindeer (R.t. platyrhynchus).

Transferrin variants in sheep: separation and characterization by polyacrylamide gel electrophoresis and isoelectric focusing.

Isoelectric focusing with carrier ampholytes in ultrathin polyacrylamide gels and polyacrylamide gel electrophoresis in a discontinuous buffer system were used for the separation of sheep transferrin variants. For identification of the different iron-binding sites of transferrin a stepwise urea gradient, different degrees of iron saturation and double one-dimensional electrophoresis were used. Isoelectric focusing results in an increased resolution of the Fe0-transferrin, Fe1-transferrin and Fe2-transferrin region. At the level of Fe0-transferrin and Fe1-transferrin the variants I, A, G, B, C, D, M, E, Q, P can be identified. The method is especially suitable for genetic studies. For screening purposes up to 108 samples can be separated within one run in an ultrathin gel.

Identification of a mouse serum protein increased by administration of an antitumor polysaccharide, PSK, as a variant of mouse transferrin and some of its biological activities.

One of the so-called LC components, the content of which is increased in the serum of PSK (antitumor polysaccharide) treated mice, was purified by repeated ion-exchange column chromatography on DEAE-Sephadex A-50. The purified protein, designated as LC-2, was identified as a variant of mouse transferrin, which is a serum beta 1-globulin having an iron-binding capacity. The absorption spectrum and ultraviolet circular dichroism (CD) curve of LC-2 were identical with those of mouse transferrin. The molecular weight, isoelectric point and amino acid composition of LC-2 were in good agreement with the results obtained previously for mouse transferrin. However, the carbohydrate content of LC-2 was different from that of mouse transferrin. This protein restored the depressed spleen cell response of tumor-bearing mice to concanavalin A (Con A) and promoted the metabolism of proteose-peptone induced peritoneal macrophages obtained from tumor-bearing mice. Furthermore, it had weak but definite antitumor activity against Sarcoma 180 cells in vivo.

Effects of Transferrin Genetic Phenotypes on Total Iron-Binding Capacity

Serum transferrin level and total iron-binding capacity (TIBC) were studied in a group of 297 healthy adult male subjects having HbAA and G6PDB+ phenotypes by standard chemical and immunoelectrophoretic techniques to examine the influence of different transferrin variants on these parameters. TIBC (67.1 +/- 13.4 mumol/l) and transferrin concentrations (27.0 +/- 3.95 mumol/l) for the whole group were found to be within reported normal values. Serum transferrin concentrations of the subjects having different electrophoretic variants and TfC subtypes of transferrin were not significantly different from each other. There was no significant difference of TIBC in relation to different electrophoretic variants of Tf. However, the TIBC of the subjects of the 1-1 subtype of TfC (70.4 +/- 13.6 mumol/l) was significantly higher than that of the 2-1 subtype (65.0 +/- 12.8 mumol/l; p less than 0.05) as well as that of the 2-2 subtype (60.4 +/- 11.4 mumol/l; p less than 0.01).