Peptide mapping of mammalian brain protein h₃in subsets of tissues and ligand binding studies

Abstract

In this report the structure of the novel polypeptide h3, isolated from subsets of tissues of a single species, and the structural similarity between interspecies h3 were investigated by peptide mapping of enzymatic and chemical cleavage fragments of h3 in one-dimensional SDS-PAGE; the peptide maps were commented on in comparison with the known sequence of 21 kDa protein, a h3-like ox brain protein. The following results were obtained: peptides generated by chymotrypsin, protease XX, BNPS-skatole and CNBr cleavage of different tissues in a single species were strikingly identical, whereas peptide maps obtained from analogue tissues in different species revealed slight structural differences. Possible ligand-h3 binding was studied by comparing the c.d. spectra of native h3, and h3 incubated with several phospholipids. Given the presence of h3 or h3-like protein in rat and human platelets, h3 was also assessed in platelet aggregation in the presence of h3 and specific anti-h3 antiserum. So far, the results emphasize the unique intra- and interspecies molecular form of h3, allow us to assign to known amino acid sequence of 21 kDa to a large extent to human h3, but do not identify h3 as a phospholipid binding protein.

Société Belge de Biochimie et de Biologie Moléculaire 162eme Réunion Antwerpen, 2 Mars 1996

“…The similar behaviour in urea-gelchromatography and urea-gradient-PAGE of the different species-h3 shows that they do not only share a common backbone structure (Bollengier et al, 1993, Seddiqi et al, 1994, but also conformational properties.…”

Amino acid sequence of the Homo sapiens brain 21-23-kDa protein (Neuropolypeptide h3), comparison with its counterparts from Rattus norvegicus and Bos taurus species, and expression of its mRNA in different tissues

“…When comparing the three mammalian proteins, three different amino acids are found in five positions: four positions are situated at the N-terminal part of the molecule (residues 19-32) and one position is located at the C-terminal region (residue 128); thus, the N-terminal part is the one most variable. Considering that we were able to demonstrate phosphatidylethanolamine-binding property for B. taurus 21-kDa protein (Bernier et al 1986), but not for H. sapiens h3 (Bollengier et al 1993), one might speculate that the observed N-terminal variability could account for functional differences between the members of the 21-23-kDa family. On the contrary, the region constituted by residues 60-126 appears to be better conserved than the N-and C-terminal sequences.…”

“…By immunoblotting, h3 was encountered in brain but also in muscle and liver from Homo sapiens, Bos taurus, Rattus norvegicus, and Gallus gallus (Bollengier et al 1983;Bollengier and Mahler 1988). Moreover, h3 isolated from different tissues in a single species appeared to be identical, whereas it revealed slight structural differences when purified from analogous tissues in different species (Bollengier et al 1993).…”