The major metabolite of dipeptide piracetam analogue GVS-111 in rat brain and its similarity to endogenous neuropeptide cyclo-L-prolylglycine.

The metabolism of a new piracetam analogue, the dipeptide cognitive enhancer N-phenylacetyl-L-prolylglycine ethyl ester (GVS-111) was studied in vivo. GVS-111 itself was not found in rat brain 1 h after 5 mg/kg i.p. administration up to limit of detection (LOD) under high performance liquid chromatography (HPLC) conditions. Three substances corresponding to the three possible GVS-111 metabolites, namely phenylacetic acid, prolylglycine and cyclo-prolylglycine, were found in experimental rat brain samples as well as in controls using HPLC, gas chromatography (GC) and gas chromatography-mass spectrometry (GC-MS) methods. Only cyclo-prolylglycine concentration increased (2.5-fold) 1 h after GVS-111 administration. Cyclo-prolylglycine formation from GVS-111 in the presence of plasma and brain enzymes was shown in vitro. These data could be considered as evidence that GVS-111 is prodrug which converts in the body to cyclo-prolylglycine, and which is identical to the endogenous cyclopeptide that produces the nootropic activity.

Amino acid sequence pattern in the regulatory peptides.

The essential properties of the primary structure of regulatory peptides, i.e. amino acid residues and their combinations, which are characteristic of the whole population of regulatory peptides, have been revealed using statistical methodology. These properties are as follows: increased content of certain residues (Gly, Pro, Phe, Arg, Tyr, Met and Trp) as well as an increased rate of occurrence of certain pairs of residue as compared with proteins, a random sequence of residues and "nonregulatory" peptides. By representing regulatory peptides as a sequence of hydrophobic (2 types) and hydrophilic (3 types) segments, the pattern for alternation of these segments in regulatory peptides has been determined. The segments were classified into 5 types according to the peculiarities of mutual localization of hydrophobic and hydrophilic residues within the primary structure of regulatory peptides. As compared with proteins, "nonregulatory" peptides and a random sequence of segments, regulatory peptides were characterized by an increased frequency of 4 particular pairs of segments among 12 theoretically possible pairs. These 4 pairs are fragments of the periodic segment sequence with periods of 4 segments. The revealed pattern indicates that there exists a general principle of the regulatory peptide primary structure organization and possibly a common type of the regulatory peptides flexible peptide chain folding at the ligand-receptor complex formation.

Phenylboronic acid as a ligand for biospecific chromatography of serine proteinases.

Via attachment of p-(omega-aminoethyl)phenylboronic acid to CH-Sepharose in the presence of water-soluble carbodiimide, a new sorbent for the biospecific chromatography of serine proteinases was obtained. The sorbent was shown to be suitable for the purification of subtilisn, alpha-chymotrypsin and trupsin. It is assumed that the serine hydroxyl group at the active site of the enzyme forms, with the boronic acid moiety of the ligand, a structure that imitates transition enzyme--substrage complex. The presence of glycerol selectively improves the binding of serine proteinases, presumably because of stabilization of the tetrahedral state of the boron atom. Direct isolation of subtilisin from a Bacillus subtilis cultural filtrate on phenylboronic acid-containing sorbent gives a virtually homogenous enzyme (42-fold purification) in a nearly-quantitative yield.