Agnes Kenessey

Effect of bacitracin on the biodegradation of β-endorphin

Abstract β-endorphin was incubated with rat brain homogenate, and the amino acids released were measured by amino acid analysis. Phe, Leu, Tyr, and Lys were liberated in the greatest amount indicating that the cleavage of Leu77-Phe78 and some Lys-X peptide bonds with endopeptidases followed by the removal of the terminal residues by exopeptidases are the main routes of β-endorphin degradation in the brain. Bacitracin considerably reduced the amino acid release from β-endorphin incubated with rat brain homogenate, and its action is suggested to be due to the inhibition of brain amino- and carboxypeptidases. Bacitracin also potentiated and prolonged the in vivo analgesic activity of β-endorphin.

STRUCTURE-FUNCTION RELATIONSHIPS IN LIPOTROPINS

Twelve years ago Li and co-workers’, 2 reported the discovery of a new lipolytic polypeptide of adenohypophysis. Because of the considerable fatmobilizing effect of this polypeptide it was designated as lipotropic hormone (LPH) or 1ipotropin.l The lipolytic activity however, has not been a specific property of lipotropin; on the contrary almost all the adenohypophyseal hormones have lipolytic side-effect in addition to their main biologic function. This fact may, at least in part, explain why comparatively less attention has been paid to lipotropin in the last ten years. During this period of time only Dr. C. H. Li’s laboratory and few additional ones, namely the authors’, Dr. M. Chretien’s, Dr. Y. Pankov’s and more recently Dr. P. J. Lowry’s group, made efforts to elucidate the structure and to understand the physiologic role of these curious polypeptides. After the discovery of enkephalins-two brain pentapeptides structurally related to lipotropin-at the end of 1975,s lipotropin, this so far “useless” polypeptide, suddenly became the focus of interest and excitement, and in turn, the knowledge of the chemistry and biology of lipotropins acquired a special value. This paper reviews the most important results achieved in our institute during the last few years, but mostly in 1976, as regards the relationships between the structure and different biologic functions of lipotropins.

Is there correlation between analgesic potency and biodegradation of enkephalin analogs

Abstract Met-enkephalin and its Pro 5 analogs containing Gly or D amino acids at position 2 were subjected to digestion with aminopeptidase M, rat brain extracts and human sera. The enzyme resistance of these peptides was compared with their analgesic activity determined in tail flick test after central and intravenous administrations. Our data did not reveal an unambigous correlation between the analgesic potency and metabolic stability of the analogs. This suggest that analgesic activity of synthetic peptides should be due to factors other than enzyme resistance /e.g. receptor binding and transport properties/.

Demonstration of β-lipotropin activating enzyme in porcine pituitary

Abstract Porcine β-lipotropin was incubated with a crude porcine pituitary homogenate, and the main cleavage products of the hormone were isolated and identified. Our results gave evidence for the enzymic cleavage of the Lys 46 -Met 47 , Arg 60 -Tyr 61 , Leu 77 -Phe 78 , and Lys 79 -Asn 80 bonds of the β-lipotropin structure. The cleavage of the Arg 60 -Tyr 61 peptide bond was accompanied with the concomitant release of opiate activity in the first period of incubation, provided that bacitracin was present in the incubation mixture. The enzyme was differentiated from trypsin or plasmin and appears to be a specific intracellular protease involved in the biosynthesis of pituitary endorphins.

Regional distribution of β-lipotropin converting enzymes in rat pituitary and brain

Among the brain areas studied only pars distalis and pars intermedia are found to contain beta-lipotropin activating enzyme indicating the these may be the exclusive organs for a physiologically significant conversion of beta-lipotropin into beta-endorphin. beta-Endorphin inactivating enzyme is found to be rather uniformly distributed in all the pituitary and brain regions. alpha-and gamma-endorphins are presumably formed by the action of the enzyme on beta-endorphin.

Trypsin-like activity of rat anterior pituitary in relation to secretory activity.

Abstract Biochemical changes in trypsin-like activity were studied in rat anterior pituitary during experimentally induced changes of hormone secretion. While dexamethasone and ACTH treatment of female rats led to a significant decrease of adenohypophyseal trypsin-like activity, elevated enzyme activity was observed after adrenalectomy and ovariectomy. This correlation together with our previous data on the subcellular localisation and some biochemical properties of the trypsin-like proteinases in the anterior pituitary suggests that these enzymes may be involved in the biosynthesis of some polypeptide hormones.

Proteolytic processing in the biosynthesis and metabolism of endorphins

All the brain and pituitary endorphins except leu-enkephalin identified to date appear to be s-lipotropin (s-LPH) fragments (Hughes et al., 1975; Guillemin et al., 1976; Li and Chung, 1976; Bradbury et al., 1976; Graf et al., 1976b ). Their structural relationships (figure 16.1) have stimulated our interest in the biosynthetic origin of these peptides. Theoretically, the release of s-endorphin from s-LPH requires a trypsin-like enzyme to split the molecule at Arg60-Tyr61. Among the naturally occurring endorphins, (s-endorphin is the most potent, implying that it may be of special physiological significance in brain function. Metenkephalin and pituitary endorphins of intermediate size might be formed from s-endorphin by further enzymatic cleavage. Their weak and transient effects in vivo give the impression that these opioid peptides may be intermediates in s-endorphin metabolism.

Interaction of 2,3-benzodiazepines with peripheral benzodiazepine receptors

2,3-Benzodiazepines (BZs), such as tofizopam (TP) and GYKI-51 189 have anxiolytic potency accompanied by moderate sedative action, but no anticonvulsant and muscle relaxant activities. These compounds show relatively low affinity to the peripheral benzodiazepine (PBZ) receptors, nevertheless, they decrease the binding of (3H)Ro5-4864 to its receptors in heart, kidney and brain membranes. This diminution in the binding is due to a decrease in the affinity for the ligand (Kd) without any change in the maximal number of binding sites (Bmax). This interaction of 2,3-BZs with PBZ binding sites may explain their pharmacological profile.