Three new genetic variants of human serum albumin have been detected in Italy by routine clinical electrophoresis. Albumin Milano Slow is common in Northern Italy, while albumins Liprizzi and Trieste, which are fast migrating, are rare and local variants. Isoelectric focusing analysis of the CNBr fragments obtained from the carboxymethylated alloalbumins in all cases localized the mutation to fragment CB5 (residues 330-446). The modified CNBr fragments were isolated on a preparative scale and subjected to tryptic digestion. Sequence determination of the abnormal tryptic peptides revealed that all the variants are caused by single point mutations: Trieste, Lys359-->Asn, Milano Slow, Asp375-->His, and Liprizzi, Arg410-->Cys. These results were confirmed by sequence determination of a variant V8 peptide in the case of Trieste, and by DNA sequence analysis for the other two variants. The DNA analysis showed a G-->C transversion at nucleotide position 11969 for albumin Milano Slow, and a C-->T transition at position 13251 for Liprizzi. The latter represents a mutation at a hypermutable CpG dinucleotide site. Albumins Trieste and Milano Slow, as most of the variants thus far described, have mutations involving residues on the surface of the molecule. In contrast, albumin Liprizzi represents the first example of a mutation in the most active binding pocket of the molecule, placed in subdomain IIIA.

A long-term electrophoretic survey on plasma proteins, which was carried out in several clinical laboratories in Italy, identified 28 different genetic variants of human serum albumin and four cases of analbuminemia. We have previously characterized 16 point mutations, 3 C-terminal mutants, and the genetic defects in two analbuminemic subjects. Here, we report the molecular defects of four alloalbumins that have been characterized by protein structural analysis. Of these, three represent new single-point mutations : albumins Tregasio, Va1122-Glu, Bergamo, Asp314+Gly, and Maddaloni, Va1.533-Met. The fourth, albumin Besana Brianza, has the same Asp494-Asn mutation that introduces a glycosylation site which has been previously reported in a variant from New Zealand, albumin Casebrook. However, in contrast to albumin Casebrook, albumin Besana Brianza is only partially glycosylated and the oligosaccharide is heterogeneous, consisting of a biantennary complex type N-glycan with either two or one sialic acid residue(s) on the antennae. Both albumin Maddaloni and Besana Brianza represent mutations at hypermutable CpG dinucleotide sites; albumin Maddaloni is a mutant that does not involve a charged amino acid.Keywords: human serum albumin; genetic variant; amino acid sequence; mass spectrometry; glycosylated alloalbumin.Alloalbuminemia is an inherited condition, which is clinically harmless, caused by a structural variant of human serum albumin. More than 100 allotypes have been found worldwide during screening of plasma proteins by routine clinical electrophoresis or in population genetics surveys with a frequency in the average population of 3.0X10-4-1.0X10~3 [I]. Many of the allotypes have been studied structurally as markers of neutral molecular evolution and because of interest in their frequency, population distribution, and ligand-binding properties. A systematic study carried out in several laboratories by protein and/ or DNA sequencing has so far identified in the albumin gene 49 single-point changes, four chain-termination mutants, and five defects causing analbuminemia (see Table 4 this involves a CpG dinucleotide in the gene and gives rise to a glycosylation site. The variant bears the same complex biantennary structure found in two other glycosylated alloalbumins, albumins Casebrook [I61 and Dalakarlia [17], but also a minor monosialylated form is present. In contrast to albumin Casebrook, albumin Besana Brianza is only partially glycosylated, the ratio between the glycosylated and unglycosylated forms being about 4: 1. MATERIALS AND METHODSElectrophoretic survey. Fresh specimens of serum from individuals with albumin Bergamo, Besana Brianza, Maddaloni, and Tregasio traits were supplied by Roberto Cesati. Each variant was named according to its geographical origin. All the donors were heterozygous and inheritance of the traits had been demonstated. A preliminary screening of the variants was per-

Metergoline, a prolactin (PRL)-lowering drug with an antiserotoninergic activity, is known to restore menstruations and fertility in hyper-PRL patients even when PRL levels are not normalized. This suggests that metergoline might also affect gonadotropins release. In a double-blind cross-over study in 8 normal males, repeated administration of metergoline enhanced the LH response to LHRH and reduced the PRL response to TRH; for females, three different tests were performed on days 5, 8 and 21 of two different menstrual cycles, each test being preceded by metergoline or by placebo administration. Metergoline always reduced the PRL response to TRH; on the 5th day, metergoline reduced the FSH response to LHRH and on the 21st day enhanced the LH response to LHRH. Basal levels of LH, FSH, T3, T4, Testosterone, 17 beta-estradiol and progesterone as well as the FSH response to LHRH (in males) and the TSH response to TRH (in both males and females) were not modified by metergoline. The data suggest that tests with TRH and LHRH can yield different results when performed during metergoline administration and that metergoline, acting through an unknown mechanism, can modify gonadotropins release.

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