Relationships Between Molecular Interactions (Nucleotides, Lipids and Proteins) and Structural Features of the Bovine Brain 21‐kDa Protein

Abstract

A cytosolic 21-kDa protein isolated from bovine brain was demonstrated to bind hydrophobic ligands, particularly phosphatidylethanolamine. It was encountered in several tissues and species ; however, its accurate function remained partially unknown. In order to obtain information from its structural features, we built a molecular model which revealed it to possess a nucleotide-binding site. In the present research, we describe the affinity of the bovine brain 21-kDa protein for nucleotides, and its association with cytosolic proteins, small GTP-binding proteins and lipid droplets. Our results suggest that, through its association with small GTP-binding proteins, the 21 -kDa protein is implicated in signal mechanisms during cell growth and maturation.From bovine brain, Bernier and Jolli% purified, in 1984, a soluble basic 23-kDa protein which showed a significant affinity for organic anions [l]. The protein was further characterized as a phosphatidylethanolamine-binding protein, suggesting its involvement in lipid metabolism [2]. The determination of its complete amino acid sequence 131 revealed the correct molecular mass to be 21-kDa. The comparison of the primary structure of the 21-kDa protein with sequence protein data banks showed very few significant sequence similarities; it was concluded that, from a structural point of view, the 21-kDa protein might represent a new type of protein [3]. Immunochemical studies using a rabbit antiserum demonstrated the presence of the 21-kDa protein in the entire cytoplasm, along the plasma membrane and the membranes of rough endoplasmic reticulum in rat brain oligodendrocytes Independent of these experiments, Bollengier and Mahler purified, in 1980, a cytosolic protein from human brain, identified as h3 [5]. In SDSPAGE, the human h3 behaved as a protein of molecular mass approximately 23 kDa. Further physicochemical studies showed that the human brain protein h3 possessed two free sulphydryl groups responsible for doublet (21 kDa and 23 kDa) and polymer formation by disulphide bonding [6]. Immunoblotting revealed the presence of protein h3 in different tissues from human, bovine, rat and chicken species 171. Recently, the human protein h3 primary structure was established both by deduction from the corresponding cDNA sequence and by direct microsequencing of the purified protein [8]; it was shown to be constituted by 186 amino acids corresponding to a calculated molecular mass of 21 kDa and present 95% sequence identity with the bovine 21-kDa protein. Furthermore, by morphine affinity chromatography, Grandy et al. 191 isolated a 23-kDa protein from rat brain membranes; although not an opioid receptor itself, the rat brain 23-kDa protein may be associated with such a receptor. The authors screened a rat brain cDNA library and established the sequence of a cDNA coding for the 23-kDa rat brain protein. The deduced 23-kDa protein shared 85% identity with the bovine 21-kDa protein and 85.5% identity with the human protein h3, revealing that the three mammalian proteins are mem...

From structure to function: possible biological roles of a new widespread protein family binding hydrophobic ligands and displaying a nucleotide binding site

“…1); interestingly, this central region corresponds to the potential nucleotide binding site observed in the bovine protein model [18]. The nucleotide site may be implicated in the binding of the bovine brain 21 23 kDa protein to small GTP-binding proteins [19]; the possibility for the mammalian protein to bind small G proteins is in accordance with the observation that TFS1 seems to be implicated in pathway modulations involving small GTPbinding proteins.…”

“…The 21 23 kDa protein was shown to associate with cytosolic proteins and small GTP-binding proteins. Our results suggested that through its association to small GTP-binding proteins, the 21-23 kDa protein could be implicated in signal mechanisms during cell growth and maturation [19].…”

“…This model contained a consensus sequence and three-dimensional folding corresponding to a potential nucleotide binding site [18]. The binding of nucleotides (especially FMN and GTP) to the bovine brain 21-23 kDa protein was then demonstrated by affinity chromatography [19]. The 21 23 kDa protein was shown to associate with cytosolic proteins and small GTP-binding proteins.…”

The Phosphatidylethanolamine-binding Protein Is the Prototype of a Novel Family of Serine Protease Inhibitors

“…Binding studies revealed its affinity for phosphatidylethanolamine (21), nucleotides like GTP and GDP and small GTP-binding proteins and other hydrophobic ligands (22). Independently, PEBP was purified from human brain as neuropolypeptide h3 (23); sequencing of this protein (24) showed 95% amino acid sequence identity with the sequence for the bovine PEBP (25).…”

NMR structure of a phosphatidyl‐ethanolamine binding protein from Drosophila

“…In mammals, bovine PEBP (bPEBP) was initially characterized as a phospholipid binding protein, 2 but it has also been shown to bind nucleotides and opioids. 3 The most abundant isoform in human, PEBP-1 (hPEBP-1/RKIP), has been described to modulate important cell mechanisms such as the control of heterotrimeric G-proteins, 4 the inhibition of mitogen-actived protein-kinase, 5 nuclear factor-kappa B signaling pathways 6 and of serine proteases thrombin, neuropsin, and chymotrypsin. 7 Human PEBP-1 is implicated in metastasis formation 8 and Alzheimer's disease.…”