the mutants of the vitamin-d-binding protein: more than 120 variants of the gc/dbp system

Vox Sang. 54: 215-225 (1988) 0 1988 S. Karger AG, Basel 0042-9007/88/0544-02 15 $2.75/0

The Mutants of the Vitamin-D-Binding Protein: More than 120 Variants of the GC/DBP System

Hartwig Clevea, Jacques Constansby aInstitut fur Anthropologie und Humangenetik der Universitat Miinchen, BRD; bCentre National de la Recherche Scientifique, Centre d‘Htmotypologie, Toulouse, France

Abstract. In this report 40 newly observed GC/DBP mutants are described. A list of the thus far identified GC mutants is presented: in addition to the three common alleles, a total of 124 Gc variants are recorded. Their population distribution is described and their relationship to the molecular features of the DBP protein is discussed. The methods currently in use for the delineation of GC mutants are briefly considered.

Introduction

The polymorphic system, known since 1959 as group- specific component (GC) [Hirschfeld, 19591, was identified by Daiger et al. [1975] as the vitamin-D-binding protein (DBP) of human plasma. DBP has been demonstrated in several mammalian species and has also been shown in fishes. It may, presumably, be present in all vertebrates as a carrier protein for vitamin D and its natural derivatives [Constans, 19841.

GC types were originally classified by immunoelectro- phoresis [Grabar and Williams, 19531. Variant phenotypes of the GC system were demonstrated by starch gel electrophoresis [Cleve et al., 19631, by a modified two-

I With the collaboration of D. Dykes (The Minneapolis War Memorial Blood Bank, Minneapolis, Minnesota, USA); C. Ehnholm (Kansanterveyslaitos Folkhalsoinstitutet, 00280 Helsinki 28, Fin- land); H. W. Goedde (Institut fur Humangenetik der Universitat Hamburg, FRG); J. Henke (Laboratorium fur forensische Blutgrup- penkunde, Diisseldorf, FRG); P. Kiihnl (Institut fur Immunhamato- logie der Universitat Frankfurt, FRG); L. L. Lai (Centre National de la Recherche Scientifique, Centre d’HBmotypologie, Toulouse, France); K. Omoto (Department of Anthropology, University of Tokyo, Japan); M. de Pancorbo (Departamento de Biologia cellular e Histologia, Universidad des Pais Vasco, Leioa, Espania); E. Simeoni (Abteilung fur Rechtsmedizin der Universitat Ee l , FRG); S. Weidinger (Institut fur Anthropologie und Humangenetik der Universitat Miinchen, FRG), and I. Yuasa (Department of Legal Medicine, Tottori University School of Medicine, Yonago, Japan).

dimensional antigen-antibody crossed gel electrophoresis [Laurell, 1965; Constans et al., 19781 and by agarose gel electrophoresis combined with immunofixation [John- son et al., 19751. The introduction of isoelectric focusing for analysis of the GC system permitted the discovery of subtypes of the GC*1 allele [Constans and Viau, 19771. The procedures currently employed are briefly described in the Methods section.

With the development of various methods for GC classification, an increasing number of common and rare inherited variants have been recognized. They were described in a series of reports: in 1979, a total of 30 GC variants was documented [Constans et al., 19791. In 198 1, the count of GC mutants was raised to 36 [Cleve et al., 198 11. In 1983, as many as 87 different GC/DBP mutants were known [Constans et al., 19831.

In this report, the presently available information on the common and rare GC alleles is presented. In addition to the three common GC alleles, a total of 124 GC mutants have been identified. Furthermore, one deficiency mutant has been recognized as well as several cases in which the existence of a silent gene GC*O has been considered [for discussion, see Vavrusa et al., 19831.

Methods

The methods for the delineation of the common and rare GC mutants have been described in detail. In this report, they are outlined only briefly.

216 Cleve/Constans

Table I. Modification of the method of Cleve et al. [ 19821.

Immobilines pK 4.6 pK 6.2

Acrylamide + Bis (28.8 + 1.2% wlv) Glycerol H2O dest. add to a total volume of PH

Acid solution Basic solution

500 p1 600 p1 300 p1 600 p1

1.3 ml 1.3 ml 2.5 g -

lOml I0 ml 4.70 5.60

PAGIF. Routine classification of GC subtypes is carried out by isoelectric focusing on horizontal flat-bed polyacrylamide gels followed by immunoprint with monospecific GC antisera [Constans et al., 19791.

For the identification of the rare GC variants additional methods are required.

PAGE. Vertical or horizontal flat-bed polyacrylamide gel electrophoresis is carried out with a Tris-borate buffer system at pH 8.6; gels are stained with Coomassie blue G 250 [Constans et al., 19791.

IEF in the Presence of 3 M Urea. Isoelectric focusing is performed in polyacrylamide gels prepared in the presence of 3 M urea. The pH gradient is established with carrier ampholytes. Gc mutants are visualized by immunoprinting [Constans et al., 19831.

IPG. The most recent methodological advance is the separation by an immobilized pH gradient [Cleve et al., 19821. The modification currently in use utilizes the recipe shown in table I.

Polymerization is carried out with TEMED and ammonium persulfate. The gel is prepared with a microgradient mixer, a peristal- tic pump is employed. The gels are poured into a cassette and subsequently kept in an oven at 50'C for 2 h in order to accelerate polymerization. Prior to separation, the gels are placed into a refrig- erator at 1o'C for 1 h. After removing the gels from the cassette they are used for isoelectric focusing without prewashing. After separation, GC is visualized on immunoprints; the gels are stained with Coomassie blue G 250.

Results

The nomenclature adopted at the international work- shop on the GC system held in July 1978 was used [Constans et al., 19791. Double-band variants are called GC 1 and single-band variants are called GC2. All double- band variants anodal to GC 1 S are called GC 1 A, all double-band variants cathodal to GClS are called GClC. Accordingly, the single-band variants anodal to GC2 are called GC2A, the single-band variants cathodal to GC2 are called GC2C. To the different GC mutants numbers are added following the chronological order of their discovery.

A complete list of the identified rare GC mutants is given in table 11. For the GC variants which were known prior to the adoption of the above outlined nomenclature, the old designation is given. Mentioned is, furthermore, in which population they were observed and whether information as to their mode of inheritance as products of alleles at the GC gene locus is available or not. Reference is given only to the summarizing reports of 1979, 198 1 and 1983 and to some original communications. For the GC mutants described for the first time in this report, the investigator and the identification of the specimen are recorded.

GClA Variants Ten newly observed 1A variants are included in table I,

which brings the number of 1A variants to a total of 32. In several instances, their IEF pattern was indistinguishable fromeitherGClF(lA24, lA26, 1A30) orGClAl (lA23, 1A28, 1A29) (fig. 1). Analysis by PAGE or by IEF in the presence of 3 M urea permits their classification, except for lA23, 1A28 and 1A30 (fig. 2,3). These were shown to be different by IPG.

GCl C Variants Sixteen additional 1 C variants have been observed.

The isoelectric points of the double bands of GClS are 4.85 and 4.95, respectively. Variant types range from these PI values to 4.98 and 6.10, respectively, as observed for GClC39. The IEF patterns of several 1C variants are very similar and, in fact, sometimes indistinguishable (fig. 1). Their differences become apparent only after analysis by PAGE and/or IEF in the presence of 3 M urea (fig. 2,3). The variant most difficult to classify was 1C38. The PAGE pattern of lC5 1 shows a more anodal electrophoretic mobility than the GClF and IS proteins. This was found previously also for lC1, 1C26 and 1C27, respectively.

GC2 Variants Six new 2A variants and eight new 2C variants have

been added to the list of single band GC mutants (table 11). Their patterns are presented in figures 1-3. It may be noted that the variants 2A5 has by IEF analysis in the presence of 3 M urea a cathodal position in comparison with GC2.

In all, three common and 124 rare GC variants have been tabulated. We mention also the deficiency mutant due the allele GC*le [Vavrusa et al., 19831, as well as the instances of a possible 'silent' gene which were observed by several investigators [Prokop and Rackwitz, 1968;

The Mutants of the Vitamin-D-Binding Protein 217

Table 11. Summary of mutants of the 6C/DBP system

Gc mutants Laboratory (Ref.) Old nomenclature geographical or ethnic origin of samples Familial transmission

1Al

1 A2 1 A3

1 A4 1 A5 1 A6 1 A7 1 A8 I A9

lAlO 1All lA12 1A13 lA14 1A15 1A16 1A17 1A18 IA19 1 A20 1A21 1A22 lA23 1A24 1A25 1A26 1A27 1A28 1A29 1A30 1A31 1 A32 1c1 1 c 2 1 C3 1 C4 1C5 1 C6 1 C7

1 C8 1 C9 lCl0 1c11 1c12 1C13 1C14 1c15 1C16 1C17 1C18

Constans et al., 1979

Constans et al., 1979 Constans et al., 1979

Constans et al., 1979 Constans et al., 1979 Constans et al., 1979 Constans et al., 1979 Constans et al., 1979 Constans et al., 1979

Cleve et al., 1981 Cleve et al., 1981 Cleve et al., 198 1 Cleve et al., 1981 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 J. Constans, 7445 J. Constans, 184-302 D. Dykes, 12063 H. W. Goedde J. Constans, 8899 J. Constans, Jura 107 LYuasa, NZ650 D. Dykes, 18 100 S. Weidinger, Glashauser J. Henke, Corsten Constans et al., 1979 Constans et al., 1979 Constans et al., 1979 Constans et al., 1979 Constans et al., 1979 Constans et al., 1979 Constans et al., 1979

Constans et al., 1979 Constans et al., 1979 Constans et al., 1979 Cleve et al., 198 1 Dykes and Polesky, 1982 Thymann et al., 1982 Thymann et al., 1982 Constans et al., 1983 Constans et a]., 1983 Constans et al., 1983 Constans et al., 1983

GcAb

GcJ GcN

GcIgl GcV2 GclC GcOp2 GcAml GcAm2 or GcD GcChip

GcT OMS 1 GcV 1 OMS2 1

GclDl GclD2 or GcOp3 GclE GcSi GclG

Australia (Aborigines), New Guinea (Aborigines), African and American Blacks, Polynesia, Indonesia Japan, Korea, China, Indonesia, Eskimos Philippines (Negritos), Japan, China, Korea, Eskimos, North American Indians Peru-Bolivia, Greenland, Canada (Eskimo), Japan Germany Djibouti (Afar-Issa), Madagascar, Ethiopia Czechoslovakia Bolivia-Peru, Asia Bolivia-Peru, Eskimos, Asia, Germany

US (Chippewa Indians), Austria Belgium, France, Germany Germany Turkey South-India (Paniyan) France Denmark Switzerland Switzerland Switzerland Germany Aborigine- Australia Bima Id., Indonesia France, Spain Mali-Bobo, Madagascar US-African-Madagascar Indian-South America, Atacaman France France, Germany New Zealand-mixed group, European-Maori Amerindian Germany Germany France, Germany, Austria, Czechoslovakia Japan, Australian Aborigines Germany, France, North and Central Africa Japan France France (Savoie) Tibet, Brazil, Czechoslovakia

France, Germany Denmark Niger, North American Indians France US (Caucasian) US (Caucasian) Denmark Germany US (Black) Japan Japan

Yes

Yes Yes

Yes Yes Yes Yes Yes Yes

yes Yes no information no information no information no information Yes no information Yes no information no information no information no information no information Yes no information Yes no information Yes no information Yes yes yes yes Yes yes Yes no information no information Yes

yes yes no information no information yes yes Yes no information no information no information no information

Cleve/Constans 218

Table 11. (cont.)

Gc mutants Laboratory (Ref.) Old nomenclature geographical or ethnic origin of samples Familial transmission

1C19 1 c 2 0 1c21 1c22 1 C23 1 C24 1C25 1 C26 1C27 1 C28 1 C29 1C30 1C31 1 C32 lC33 1C34 1C35 1 C36 IC37 1 C38 1 c39 1 C40 1C41 1 C42 1 C43 1C44 lC45 1 C46 1 C47 1 C48 1 C49 1C50 1C51 IC52 1C53

Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et a]., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 M. Pancorbo, Bask 1 18 D. Dykes, 14788 M D. Dykes, 12846 AF D.Dykes, 12695 D. Dykes, 14032 Constans, 8904 J. Constans, I86/80 L. L. Lai, Aust. 1640 D.Dykes 14969 M H. Cleve, A. L. P. Kiihnl, E. Simeoni D. Dykes, 20095 AF K. Omoto, 8-29 H. Cleve, lsr. 1 129 1 D. Dykes, 2331 8 AF D.Dykes, 21817C

France France, Canada France Denmark US (Black) Australia (Aborigines), Indonesia (Timor) US (Caucasian) France Germany Bali (Chinese) Bali (Chinese) Bali (Tinadja) US (Caucasian) US (Caucasian) US (Caucasian) Gambia Japan, Australian Aborigines Nepal Buka (Island) (Bougainville) Basque (Spain) US (Caucasian) US (Caucasian) US (Caucasian) US (Black) France Madagascar Australia US (Mexican) Germany Germany US (Black) China Israel US (Caucasian) US (Black)

no information no information no information no information no information Yes Yes no information Yes no information no information no information Yes no information no information no information Yes Yes Yes no information no information no information no information no information no information no information no information no information

Yes no information no information no information no information no information

yes

2A 1 2A2 2A3

2A4 2A5 2A6 2A7 2A8 2A9 2A10 2A11 2A12 2A13 2A14 2A15 2A16 2A17

Constans et al., 1979 Gc2C Constans et al., 1979 GcOpl Constans et al., 1979 Gc2A

Constans et al., 1979 GcY Constans et al., 1979 Constans et al., 1979 GcWien Cleve et al., 198 1 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 D.Dykes, 41693 D. Dykes, 18807 C D. Dykes, 11622

Gc2B, Gcv3

West Africa (Niger) Germany, Czechoslovakia, Sweden West-Central Africa (Pygmies), US (Black), Madagascar (Mala- gasy), Australia (Aborigines) Japan Senegal-Gambia, US (Black), Israel (Druzes) Austria, US (Caucasian) Japan Germ any Scotland, Sweden, Denmark Guam (Chamorros) Japan US (Caucasian) US (Caucasian) Japan US (Caucasian) US (Asia) US (Caucasian)

no information Yes Yes

Yes Yes Yes no information no information Yes no information Yes no information yes yes no information no information no information


Table 11. (cont.)

Gc mutants Laboratory (Ref.) Old nomenclature geographical or ethnic origin of samples

2A18 2A19 2A20 2 c 1 2c2 2C3 2C4 2C5 2C6 2C7 2CS 2C9 2CIO 2c11 2c12 2C13

2C14 2C15 2C16 2C17 2C18 2C19

C. Ehnholm, S. P. K.Omoto, I 176 K. Omoto, I 59 Constans et al., 1979 Gc2D Constans et al., 1979 GcZ Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 Constans et al., 1983 H. Cleve, U. J. 1. Yuasa Gc2C

D.Dykes, JMC D. Dykes, 14593 1.Yuasa NZ654 H.Cleve, Ma. S. Weidinger, Metz. Ch. D.Dykes, 21377C

Auckland

Finland Japan Japan France Germany, Denmark Germany Armenian Sweden Iranian US (Caucasian) France Bali (Chinese) Switzerland US (Caucasian) Germany New Zealand

US (Caucasian) US (Caucasian) New-Zealand-European-Maori mixed group Germany Germany US (Caucasian)

Familial transmission

no information no information no information Yes Yes no information no information Yes Yes Yes no information no information no information no information no information no information

no information no information no information Yes no information no information

Patscheider and Dirnhofer, 1979; Brinkmann et al., 1981; Suzuki et al., 19861.

The distribution of the rare GC variants is given in table 111. 85 (68.6%) represent double-band mutants; 39 (31.4%) are single-band mutants. Among the GC1 mutants the cathodal variants are prominent. The GC2 mutants are equally distributed in the anodal and in the cathodal group, respectively.

The geographical or ethnical distribution is summa- rized in table IV. This distribution reflects the intensity of examination of the GC system in the various parts of the world. Thus, most of the variants were found in Europe, in North America and Japan. Although more than 160 different populations have been tested by IEF for the GC polymorphism, the largest number of individuals has been examined in Europe (approximately 12,500), North America (approximately 10,000 Caucasians and 1,200 US Blacks) and in Japan (approximately 4,000).

Some of the variants listed in table I are not infrequent in certain populations: in Europeans 1A5, 1C1, 1C3, and 2C2 were found in several surveys. In the Mongoloids, the variants 1A2,1A3 and 1A9 are fairly frequent. In African and American Blacks lA6, 1C3, 1C10,2A3 and 2A5 are

Table 111. Distribution of GC mutants ~~ ~~ _______~

GC Total GClA GClC GC2A GC2C mutants

Number 124 32 53 20 19 Percent 100 25.8 42.8 16.1 15.3

Table IV. Geographical and ethnic distribution of GC variants.

Population GClA GClC GC2A GC2C

Caucasians 17 32 9 14 (Europe and North America) Blacks 4 9 3 (Africa and America) Asian Populations

Near East 2 I 1 2 Far West 8 12 8 1

Eskimos 4 American Indians 7 2 Australian Aborigines 2 4 1 and Melanesians Polynesians 2 1 2

220 ClevelConstans

Fig. 1. Schematic presentation of phenotypes of the GC/DBP system obtained by isoelectric focusing on polyacrylamide gels followed by immunoprint a GC 1A variants. b GC 1C variants. c GC 2A and 2C variants.

The Mutants of the Vitamin-D-Binding Protein 22 I

Fig. 2. Schematic presentation of phenotypes of the GClDBP system observed by polyacrylamide gel electrophoresis. a GC 1A and 1C variants. b GC 2A and 2C variants.

observed with varying frequencies in different populations. Eskimos are characterized by the variant 1 A4, some American Indian tribes have 1 AS, lA9 and 1 C7 which are also encountered in Asian populations in the Far East [Constans et al., 1985; Kamboh and Ferrell, 19861.

The ‘fate’ of the two genetic variants of the GC system which were discovered first in 1963 was very interesting. GC Aborigine and GC Chippewa, now known as GC 1 A 1 and GClAlO [Cleve et al., 19631.

GC Aborigine was originally observed in Australian aborigenes, later found in Melanesians and in African and American Blacks [Kirk et al., 1963; Cleve et al., 1967; McDermid and Cleve, 19721, as well as in populations

from Malagasy and Bali [Constans et al., 19851. Speci- mens with 1Al from these different populations were examined in comparison by IEF, PAGE, IEF in the presence of 3 M urea and by two-dimensional antigen-antibody crossed gel electrophoresis. They were found to be identical and represent, thus, presumably the same wide- spread mutant.

Gc Chippewa, on the other hand, has not been found in any other American Indian tribe. GClAlO is present in the Chippewas south from the Lake Superior [Cleve et al., 19631 as well as in the Ontario Chippewas, called Ojib- was, in the north of the Great Lakes [Szathmary et al., 19741. lAlO was recently encountered in a sample from

ClevelConstans 222

Fig. 3. Schematic presentation of phenotypes of the GClDBP system analyzed by isoelectric focusing on polyacrylamide gels in the presence of 3 M urea followed by immunoprint a GC 1A variants. b GC 1C variants. c GC 2A and 2C variants.


Austria and may, thus, represent originally a ‘European’ mutant, possibly introduced into this Indian tribe in the 17th or 18th century. The high frequency for GClAlO in the Chippewa tribe may be due to a founder effect with subsequent increase of the frequency of this allele in this particular population.

Discussion

The results presented in this report emphasize the need for the application of a variety of methods for the classification of genetic variants of the DBP system as well as of proteins in general. Examination with a single method may serve only as a screening procedure. In addition to the procedures described in the Methods section, analysis by two-dimensional electrophoresis may be useful in particular cases [Vavrusa et al., 19831. IEF with immobilized pH gradients represents undoubtedly a further powerful method for the delineation of genetic variability. It has to be kept in mind, however, that in several instances shifts in the positions of certain variants have been noticed when IEF with carrier ampholytes (CA) was compared with IEF with the use of IPG [Gorg et al., 1985; I. Yuasa, D. W. Cox, respectively, pers. commun.]. The reason for this phenomenon is not clear: CA may interact with protein and form protein-CA complexes with slightly altered PI values. On the other hand, in the IPG system ionic strength is low and free electrostatic groups are present, which may influence solubility and/or mobility of proteins. At the present time, IPG, thus, represents a complementary method for the characterization of variants in particular and difficult cases.

Analysis at the protein level discloses genetic variability which affects the primary structure of the gene products. Mutations at the genic level which do not alter the amino acid sequence are not ascertained. The recent advances in nucleotide sequencing at the niveau of cDNA or genomic DNA undoubtedly added a new dimension to the analysis of genetic variability. The approaches of analysis at the gene product and at the genic level will be in the future complementary rather than exclusive. Com- parison of the cDNA sequences and their derived amino acid sequences of the DBP type GC2 with GC 1 has shown not only four amino acid substitutions as the possible allelic differences but, in addition, two base substitutions in third codon positions which are not reflected in the gene product [Yang et al., 1985; Cooke and David, 19851.

The functionally active sites of the DBP molecules, the binding site for the vitamin D and its natural derivatives as well as the binding site for actin, have not yet been definitely assigned to circumscribed portions of one of the three molecular domains.

The genetic differences range from positions 118 to 420 of the amino acid sequence. The species differences between human and rat DBP concern amino acid differences in domain I11 and in part of domain 11. A long sequence in between is preserved and may, thus, be relat- ed to the function of DBP [Schoentgen et al., 1986; Cooke, 1986; Constans, 19861. It is unlikely that the vast number of GC/DBP mutants is randomly distributed. Refined methods have to be developed to permit identification of the structural basis of these mutants.

The cause for the double-band pattern of the GC1 mutants has been established. The anodal GC isoprotein contains a trisaccharide as a carbohydrate side chain which has N-acetyl neuraminic acid (NANA) as terminal residue [Viau et al., 19831. Comparative experiments after treatment with neuraminidase revealed that the GC1 subtypes and the majority of GC1 double-band mutants have one NANA residue per GC 1 a isoprotein, while the cathodal isoprotein GClc does not contain NANA [Cleve and Patutschnick, 19791. GC2 and GC2 mutants also do not contain NANA [Cleve and Patutsch- nick, 19791. Some notable exceptions to this rule have been found: GClA16 has two NANA residues in the anodal and one NANA residue in the cathodal isoprotein. GC 1 A 1 8 and 1 A 1 1 are mutants with three and two NANA residues in the anodal and cathodal isoproteins, respectively [Thymann et al., 19851. Among the GC2 mutants, 2A3 and a single band mutant called GCAr have one, 2A14 has two NANA residues [Constans, 1981;Nakasoneetal., 1983,19851.Itmaybepointedout that the distance in the IEF pattern between the anodal and the cathodal isoprotein is maintained in all GC double-band mutants. This observation makes the presence of two structurally different polypeptide chains, apart from the 0-glycan chain differences, very unlikely.

A deficiency of the GC protein has, thus far, been observed only in the heterozygous state. These persons had GC plasma concentrations reduced to levels of one quarter to one half of the normal concentrations. These states are reportedly compatible with good health [Vav- rusa et al., 19831. None of the diseases with autosomal- recessive mode of inheritance could, thus far, be asso- ciated with quantitative or functional GC deficiency states.

ClevelConstans 224

Acknowledgements

We would like to thank Mrs. C. Gouaillard, Mr. A. Clerc (Tou- louse) and Ms. A. Brandhofer (Munich) for expert technical assis- tance.

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McDermid, E.M.; Cleve, H.: A comparison of the fast migrating Gc-variant of Australian Aborigines, New Guinean Indigenes, South African Bantu, and Black Americans. Hum. Hered. 22:

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261: 3441-3450 (1986).

241-253 (1972).

184-185 (1983).


Szathmary, E. J. E.; Cox, D. W.; Gershowitz, H.; Rucknagel, D. L.; Schanfield, M. S.: The Northern and Southeastern Ojibwa: serum proteins and red cell enzyme systems. Am. J. phys. Anthrop. 90:

Thymann, M.; Hoste, B.; Scheffrahn, W.; Constans, J.; Cleve, H.: Unusual sialilation of three different rare genetic variants of serum DBP: GclA17, GclA16 and GclAl1. Hum. Genet. 69:

Thymann, M.; Hjalmarsson, K.; Svensson, M.: Five new Gc variants detected by isoelectric focusing in agarose gel. Hum. Genet. 60:

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49-66 (1 974).

224-227 (1985).

340-343 (1982).

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Received: October 19, 1987 Accepted: November 2 , 1987

Prof. Dr. Hartwig Cleve Institut fur Anthropologie und Humangenetik der Universitat Munchen Richard-Wagner-Strasse 1 O/I D-8000 Munchen 2 (FRG)

the mutants of the vitamin-d-binding protein: more than 120 variants of the gc/dbp system

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