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K Barnes

Publications and source records attributed to K Barnes.

8 recordsLinked to original sources
PubMed1992

A comparison of the zinc contents and substrate specificities of the endothelial and testicular forms of porcine angiotensin converting enzyme and the preparation of isoenzyme-specific antisera.

Angiotensin converting enzyme (ACE; EC 3.4.15.1) was purified from porcine kidney and lung (endothelial isoenzyme) and testis (testicular isoenzyme) by affinity chromatography on lisinopril-2.8 nm-Sepharose. Atomic-absorption spectroscopy revealed that ACE purified from kidney and lung contained 2.58 and 2.35 atoms of zinc per molecule of enzyme (M(r) 147,000) respectively. In contrast, ACE purified from testis contained only 1.58 atoms of zinc per molecule of enzyme (M(r) 80,000). Thus it would appear that both putative zinc-binding sites in endothelial ACE contain zinc and may therefore be catalytically active. No differences were observed in the pattern of products generated on hydrolysis of benzoyl (Bz)-Gly-His-Leu, substance P, luteinizing-hormone-releasing hormone (LH-RH) and its analogue, des-Gly10-LH-RH-ethylamide, by kidney and testicular ACE. There was also no difference in the initial rates of hydrolysis of Bz-Gly-His-Leu or substance P by the two isoenzymes, although LH-RH and its analogue were hydrolysed twice as rapidly by kidney ACE. It is therefore unlikely that the N-terminal catalytic site in porcine endothelial ACE is predominantly responsible for the atypical cleavage of LH-RH generating the N-terminal tripeptide. Two polyclonal antisera were raised to the affinity-purified forms of pig kidney and testicular ACE. Isoenzyme-specific antisera were then isolated from these by absorbing out those antibodies recognizing determinants on the other isoenzyme. Immunoelectrophoretic blot analyses and immunofluorescent staining of sections of pig kidney were used to demonstrate the specificity of the antisera. Immunofluorescent staining of sections of pig testis with the antiserum specific to testicular ACE localized testicular ACE solely to the lumen of the seminiferous tubules, whereas the antiserum specific to endothelial ACE revealed the presence of this isoenzyme only in blood vessels. The antiserum to endothelial ACE, which recognizes determinants in the unique N-terminal domain, was investigated as a possible specific inhibitor of the N-terminal catalytic site. Although this antiserum failed to inhibit testicular ACE, the effect on the activity of endothelial ACE appeared to be due to inhibition of both the N- and C-terminal catalytic sites.

Amino Acid Sequence↗
PubMed1992

Endoscopic carpal tunnel release: an anatomic study of the two-incision method in human cadavers.

To determine the relationship of neurovascular structures to the sites of portal placement and transverse carpal ligament division during two-portal endoscopic carpal tunnel release, a study of 20 fresh cadaver specimens was carried out. Open dissection of the carpal tunnel after endoscopic surgery showed complete ligamentous release in 18 hands (90%). In 10 specimens, the procedure was performed as described by Chow. There was one partial transection of the superficial palmar arch (5%), and five specimens (50%) had complete divisions of the superficial palmar fascia with considerable pressure placed on the ulnar nerve at the wrist. A modified technique was used in 10 specimens in which the proximal incision was made in a more distal location and a distally based ligamentous flap was created. The superficial palmar arch and the distal edge of the transverse carpal ligament were visualized directly before passage of the trocar. No complications were noted with this method.

Anthropometry↗
PubMed1992

Membrane localization of endopeptidase-24.11 and peptidyl dipeptidase A (angiotensin converting enzyme) in the pig brain: a study using subcellular fractionation and electron microscopic immunocytochemistry.

Brains from piglets were dissected and a block of tissue including the substantia nigra, globus pallidus, and entopeduncular nucleus was homogenized and then fractionated on discontinuous Percoll gradients. Ligand-binding assays using (-)-[3H]nicotine and [3H]quinuclidinyl benzilate served to delineate fractions containing nicotinic and muscarinic acetylcholine receptors. In this system endopeptidase-24.11 exhibited a biphasic distribution, consistent with its presence on both pre- and postsynaptic membranes. Peptidyl dipeptidase A (angiotensin converting enzyme; ACE) was associated with membrane fractions containing muscarinic receptors. An immunoblot of these fractions with an affinity-purified polyclonal antibody to ACE revealed only the neuronal form of ACE (Mr 170,000), the endothelial form (Mr 180,000) being undetectable. Electron microscopic immunoperoxidase staining of the substantia nigra, with an affinity-purified antibody to endopeptidase-24.11 at the preembedding stage, showed this antigen to be confined to the plasma membranes of boutons, axons, and some dendrites. Both pre- and postsynaptic membranes were stained, and occasionally other regions of the dendritic membrane were positive. No staining of synaptic vesicles within the boutons was observed. Thus, two independent approaches indicate that endopeptidase-24.11 is present on both pre- and postsynaptic membranes in the pig substantia nigra. The subcellular fractionation suggests that neuronal ACE is confined to dendritic membranes.

Animals↗
PubMed1991

Membrane peptidases in the peripheral nervous system of the pig: their localization by immunohistochemistry at light and electron microscopic levels.

The presence and cellular localization of five membrane peptidases has been investigated in peripheral nerves, including those of the autonomic nervous system, in the pig. Endopeptidase-24.11 ("enkephalinase") peptidyl dipeptidase A, aminopeptidase N, aminopeptidase W and dipeptidyl peptidase IV were studied by both enzymic assays of membranes prepared from samples of nerve and by immunoperoxidase histochemistry at light and in two cases, endopeptidase-24.11 and aminopeptidase W, at electron microscopic levels. All five peptidases could be quantified by enzymic assay, though the activities were about 1% of those in renal microvilli and less than those of choroid plexus membranes. Endopeptidase-24.11 was associated with Schwann cell membranes in all types of nerve examined, including major nerves containing predominantly myelinated fibres as well as autonomic nerves, such as the vagus and splenic nerves and the sympathetic chain, staining being observed in membranes associated with myelinated and unmyelinated fibres. The Schwann cell location of endopeptidase-24.11 was confirmed by correlation with immunostaining for glial fibrillary acidic protein and by electron microscopy. This peptidase is known to have a wide repertoire of susceptible substrates among neuropeptides which was here shown to include vasoactive intestinal polypeptide (Km 268 microM, kcat 568 min-1), one of a number of neuropeptides present in peripheral nerve fibres. Three of the peptidases, peptidyl dipeptidase A, aminopeptidase N and dipeptidyl peptidase IV, were associated with microvessels of peripheral nerves. Aminopeptidase N was also observed in connective tissue elements, including the perineurium. Aminopeptidase W was unique among the five peptidases in having a neuronal localization. This was observed in unmyelinated and myelinated nerves and was supported by comparison with the pattern of staining observed for neurofilament protein and by electron microscopic immunoperoxidase staining. This observation was unexpected since aminopeptidase W has not been detected as a neuronal marker in the brain. Some possible roles for the membrane peptidases in peripheral nerves are discussed.

Animals↗