Orosomucoid (ORM) typing by isoelectric focusing: evidence for an additional duplicated ORM1 locus haplotype and close linkage of two ORM loci.

Human serum orosomucoid (ORM) exhibits a high variability. Several alleles including a duplicated ORM1 gene (ORM1*2.1) have been identified at two functional ORM loci, ORM1 and ORM2. In this study a modified isoelectric focusing, in which glycerin was omitted from gels, was used to differentiate a new variant ORM1 5 from ORM2 3. The ORM1 5 band was always observed together with the ORM1 2 band. The simultaneous expression could be explained in terms of an additional duplicated ORM1 locus haplotype, ORM1*5.2, whose average frequency was .016 in two Japanese populations. The ORM2*6 found at a polymorphic frequency (.023) was demonstrated to be in association with ORM1*2, indicating the close proximity between ORM1 and ORM2 loci.     

Orosomucoid (ORM) typing by isoelectric focusing: evidence for gene duplication of ORM1 and genetic polymorphism of ORM2

Summary It has been demonstrated that the genetic polymorphism of human serum orosomucoid (ORM) is controlled by polymorphic ORM1 and monomorphic ORM2 loci. In this study a Japanese family was encountered in which several members had puzzling electrophoretic patterns consisting of four bands. The ORM patterns were due to the products of a duplicated ORM1 locus haplotype (ORM1 ^* 2·1) or the products of new variant alleles at the ORM2 locus. The ORM1 ^* 2·1 haplotype is very common in the Japanese population, occurring at an allele frequency of 0.16. The increased occurrence of ORM1 2-1 and the heterogeneity in band intensity among ORM1 2-1 phenotypes could be explained in terms of a duplicated gene ORM1 ^* 2·1. The ORM2 locus proved to be polymorphic, with six alleles in the Japanese population. Dedicated to Professor Dr. K. Nishigami on the occasion of his 60th birthday     

Orosomucoid (ORM) typing by isoelectric focusing: an analysis of ORM haplotypes

A new separator isoelectric focusing method for typing of orosomucoid (ORM) was developed. This method provided a superior resolution of ORM patterns: two close bands of ORM1*5.2 products were clearly separated. A total of 364 subjects from Okinawa (Japan) were classified into 21 ORM phenotypes determined by 6 ORM1 and 7 ORM2 alleles including a polymorphic silent allele, ORM2*QO, and 2 new rare variants, ORM2*18 and ORM2*19. These phenotypes were also explained by 12 ORM haplotypes, half of which were polymorphic.     

Orosomucoid (ORM) typing by print lectinofixation: a new technique for isoelectric focusing. Two common alleles in Japan

Summary The genetic types of orosomucoid (ORM) were analyzed by isoelectric focusing (IEF) on polyacrylamide gels and subsequent print lectinofixation with a lectin from the beetle, allo A. In this paper, the newly devised print lectinofixation for ORM typing is described. This technique is faster, easier to perform, and has been found to be a useful tool in population genetics and forensic medicine. The results of typing for two alleles, ORM ^*1 and ORM ^*2 are described for a population of Northern Japan (n=500). We use the designation “lectinofixation” to denote the method using lectin in place of monospecific antibody in the immunofixation     

Orosomucoid (ORM 1) subtyping and formal genetics

Summary Orosomucoid (ORM) phenotyping has been performed in 141 families with 407 children from southwest Germany. Eight families were observed in which duplicated ORM1 genes, F1F2, F1F3, segregated. The family data gave no information about the presence of tandemly duplicated ORM1 F1F4 and ORM1 F1F5 genes. To date, the segregation of the phenotypes of the children agrees with the extended formal model: two ORM1 loci with two common (^*F1, ^*S) and several rare (^*F1F2, ^*F1F3, ^*F4, ^*F5) alleles. The parental allele frequencies were calculated by gene counting as ORM1 ^*F1 = 0.5781, ^*S = 0.3901, ^*F1F2 = 0.0195, ^*F4 = 0.0053, ^*F1F3 = 00.0035, ^*F5 = 0.0035.     

Three new orosomucoid (ORM) variants revealed by isoelectric focusing and print immunofixation

Summary Phenotypes of orosomucoid (ORM) in human sera have been analysed by isoelectric focusing and print immunofixation. After neuraminidase treatment the band patterns indicated that the polymorphism of the structural locus ORM1 is controlled by three autosomal codominant alleles. According to the previous nomenclature they were called ORM1^*F1, ORM1^*F2, and ORM1^*S. In a study of 272 unrelated individuals from southern Germany, five of the six expected common ORM1 subtypes were observed. Furthermore, we found three ORM variant phenotypes which have not been reported previously. These variants were characterized by additional bands in a cathodal position. One variant had additional double bands and presumably represents a rare ORM1 variant named ORM1S1. Two variants had additional single bands. They were assigned tentatively to the ORM2 gene locus. While the common gene product of ORM2 may be called ORM2A, the two variants are named ORM2B1 and ORM2B2, respectively. ORM2B1 has, thus far, been found only in a single individual; the variants ORM1S1 and ORM2B2 were found in a father-child pair and a mother-child pair, respectively. The frequency for variants tentatively assigned to the ORM2 locus is very low and was calculated to be 0.0037.     

Orosomucoid (ORM1 and ORM2) types in the Spanish Basque Country, Galicia and northern Portugal

The genetic variation of orosomucoid (ORM1 and ORM2) in three south-western European populations (Galicia, Spanish Basque Country and northern Portugal) was investigated using hybrid isoelectric focusing. Three common ORM1 alleles were observed in these populations, the frequencies of ORM1 *S observed in Galicia and northern Portugal being the highest found among populations of European origin. Rare variants were observed for both the ORM1 and ORM2 loci.     

Worldwide distribution of phosphoglucomutase 1 (PGM1) polymorphism detected by isoelectric focusing: A review

This paper reports an exhaustive and updated compilation of phenotype and allele frequency data for phosphoglucomutase locus 1 (PGM1), obtained with an analytical isoelectric focusing technique, in human populations. The analysis of the PGM1 allele frequency distributions within and among the major human groups together with the degree of diversification evaluated by Wright's F_st, computed per allele and averaged over alleles, are also presented.     

Investigations on the PGM 1 a polymorphism (phosphoglucomutase-EC 2.7.5.1) by isoelectric focusing

Summary The determination of phosphoglucomutase (PGM₁) phenotypes was performed by isoelectric focusing on samples from 1678 unrelated individuals from Hessen, Germany. Ten common phenotypes are considered as gene products of four alleles at the PGM₁ locus with the following frequencies: PGM ₁ ^a1 =0.6305, PGM ₁ ^a2 =0.1844, PGM ₁ ^a3 =0.1320, and PGM ₁ ^a4 =0.0530. Twenty-two different mating types were observed in 113 families with 202 children. The segregation of the phenotypes in the offspring supports the assumed way of autosomal codominant inheritance. The example of a silent allele (PGM ₁ ⁰ ) as well as a rare variant (PGM ₁ ⁷ ) is reported.     

I (C3b/C4b inactivator) typing by agarose gel isoelectric focusing and immunoblotting technique

Genetic polymorphism of I (C3b/C4b inactivator) was studied by the method of agarose gel isoelectric focusing followed by an immunoblotting technique. Serum or plasma samples were pretreated with neuraminidase. The method is rapid, and gives the simple and reliable patterns of I. The allele frequencies calculated from healthy Japanese individuals living in the western part of Japan were: IF* A = 0.126 and IF*B = 0.874.     

Pituitary protein lipolytic factor(s): Partial purification by isoelectric focusing (IEF)

Tests were made on highly purified bovine peptidal pituitary factors for lipolytic activity on rat adipose tissue. The lipolytic protein factor from bovine pituitary glands was isolated and characterized by ethanol precipitation, gel filtration, and isoelectric focusing. The supernatant of the homogenized tissue was precipitated with ethanol, the precipitate was lyophilized and resuspended in HCl 1 mmole/liter, the insoluble was discarded, and the supernatant lyophilized. A solution of the lyophilized fraction was fractionated by gel filtration on Sephadex G-75. The bands with lipolytic activity were pooled and lyophilized. Lipolytic activity was determined by incubation of epididimal rat adipose tissue and by successive measurements of free fatty acids and glycerol released in the incubation medium. The gel filtration with the highest lipolytic activity (20.4 µequiv/g per 4 hr and 17.0 µmole/g per 4 hr glycerol) was submitted to isoelectric focusing. The gel filtration still appeared highly heterogeneous, but most of the lipolytic activity was concentrated in the protein band at pH 8.6.     

Group-specific component (Gc) and transferrin (Tf) subtypes ascertained by isoelectric focusing

Simultaneous subtyping of Gc and TfC by isoelectric focusing allows us to compute the following gene frequencies for the Belgian population: Gc1S = 0.543 Gc1F = 0.167 Gc2 = 0.290 TfC1 = 0.784 TfC2 = 0.206 TfB = 0.007 TfD = 0.003. The Gc bands were precipitated by sulfosalicylic acid instead of by immunofixation.     

Variation in turtle myoglobins (subfamily emydinae: Testudines) examined by isoelectric focusing

• 1.1. Myoglobins from heart and skeletal muscle of turtles were analyzed by thin-layer isoelectric focusing.
• 2.2. Within the subfamily Emydinae, variation in the occurrence of two myoglobin electromorphs (pI 6.8 and 6.9) was detected.
• 3.3. Patterns of myoglobin polymorphism support dividing the Emydinae into two subfamilies and help resolve controversial theories on relationships of the genus Deirochelys.
• 4.4. Possible adaptive significance of the myoglobin variants (isoforms) remains to be determined.

     

Characterisation of the isoenzymes of phosphoglucomutase (PGM) determined by the first (PGM1) and second (PGM2) locus observed by isoelectric focusing

Summary The existence of four alleles of phosphoglucomutase (PGM₁) in human red cell lysates has previously been demonstrated by isoelectric focusing (Bark et al., 1976; Kühnl et al., 1977; Sutton and Burgess, 1978). Experiments are now described in which the position of each of the first-locus (PGM₁) and second-locus (PGM₂) isoenzymes is defined, thus extending and confirming the original proposal made by Bark et al.     

Gc subtypes identified by isoelectric focusing in Baboons (Papio hamadryas)

Summary Phenotyping for Gc variants by conventional electrophoresis in 835 Papio hamadryas baboons demonstrated a monomorphic population. Gc subtyping by polyacrylamide IEF gels, pH 4–6, on 394 of these baboons revealed the existence of two common alleles which we named Gc _Papio ¹ and Gc _Papio ² . Pedigree data confirmed the inheritance of a single locus, two allele system and the observed gene frequencies were 0.593 for Gc _Papio ¹ and 0.407 for Gc _Papio ² .     

Resolution of highly purified toxic-shock syndrome toxin 1 into two distinct proteins by isoelectric focusing

Highly purified toxic-shock syndrome toxin 1 (TSST-1) was prepared by differential precipitation with ethanol and resolubilization in water followed by successive electrofocusing in pH gradients of 3-10, 6-8, and 6.5-7.5. TSST-1, thus isolated, migrated as two distinct protein bands with isoelectric points of 7.08 (TSST-1a) and 7.22 (TSST-1b). When tested by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, both toxins migrated as homogeneous bands with molecular weights of 22 000. The gel bands were visualized by silver staining. The two toxins have nearly identical amino acid compositions and are immunologically identical as shown by Ouchterlony reactivity against TSST-1 hyperimmune serum. TSST-1a and TSST-1b have the same biological activities as TSST-1: the capacity to induce fever, enhancement of host susceptibility to lethal endotoxin shock, nonspecific T lymphocyte mitogenicity, and suppression of immunoglobulin M synthesis against sheep erythrocytes. These two proteins have been isolated from several different TSS-associated Staphylococcus aureus strains. The data suggest that the differences in isoelectric point result either from the presence of a cofactor or from alternative conformations. Since only two bands appear, microheterogeneity as a result of deamination or acetylation is unlikely.     

Population genetics of the group specific component (Gc) and phosphoglucomutase (PGM1) studied by isoelectric focusing

For the determination of the group-specific component (Gc) and phosphoglucomutase (PGM1) phenotypes, isoelectric focusing was performed on two samples, one of Jat Sikh of northwest India, the other of northeast English. The subtype frequencies of these two systems do not differentiate the two populations sampled. Synthesis of the existing data shows distinct PGM1 and Gc subtype frequencies in various ethnic and racial groups. The anthropological implication of these subtype frequencies is discussed.     

Method for the typing of the C3 polymorphism in stored sera by means of isoelectric focusing

A method is described by which the three common phenotypes of C3 can be typed by desialylation and isoelectric focusing in serum samples stored at -20 degrees C for several years.