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A G Booth

Publications and source records attributed to A G Booth.

28 records · Page 2Linked to original sources
PubMed1976

Proteins of the kidney microvillus membrane. Identification of subunits after sodium dodecylsullphate/polyacrylamide-gel electrophoresis.

The proteins of microvilli prepared from pig kidney were analysed by polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulphate. The typical pattern stained for protein revealed five major bands, four of which also stained for carbohydrate, and about 15 minor bands. For descriptive purposes the bands were designated numerically by their apparent molecular weights (X10(-3). Well-characterized proteins were identified with four of the five major bands. Dipeptidyl peptidase IV, a serine proteinase that may be specifically labelled with di-isopropyl [32P]phosphorofluoridate, was assigned to band 130. Aminopeptidase M was assigned to band 160, though when released from the membrane by a proteinase, this protein comprises three polypeptides each of lower apparent molecular weight than the native enzyme. Neutral endopeptidase can be assigned to band 95 and actin to band 42. The fifth major band (180) is an extrinsic glycoprotein that has not been identified with any microvillus enzyme activity. These four proteins contribute 21% of the microvillus-membrane protein. Kidney microvillus actin was characterized by a variety of properties and was similar to muscle actin. A computer analysis of the gel pattern indicates that it comprises 9.0% of the microvillus protein. Myosin is not present in the microvillus, but another protein associated with band 95, with properties that distinguish it from neutral endopeptidase, was tentatively identified as alpha-actinin. Alkaline phosphatase was identified as a monomeric polypeptide with an apparent molecular weight of 80000; it is a minor protein of the microvillus and is not discernible as a discrete band in the gel pattern. These and other results permit a model of the organization of the microvillus protein to be suggested. The computer program used has been deposited as Supplementary Publication SUP 50070 (12 pages) at the British Library Lending Division, Boston Spa. Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms given in Biochem. J. (1976) 153,5.

Actins↗
PubMed1976

A morphometric and biochemical investigation of the vesiculation of kidney microvilli.

The vesiculation of kidney proximal tubule microvilli has been examined in tissue slices, isolated brush borders and isolated microvilli. Vesiculation could be induced in tissue slices by 2,4-dinitrophenol and anoxia. Cycloheximide and fluoride had no effect. In brush borders and microvilli, the vesiculation was found to be essentially temperature-dependent. Whilst an osmotic swelling could be produced by hypo-osmolar media, the temperature-dependent vesiculation could not be prevented in hyper-osmolar media. Of a wide variety of reagents tested, only glutaraldehyde, mercuric chloride and mersalyl were effective in arresting the vesiculation. Electron micrographs show that vesiculation involves a collapse of the internal structure of the microvillus. However, the collapse was not associated with depolymerization of the microvillus actin filaments. Rather it appeared to be due to the parting of cross-bridges between the membrane and the actin filaments. The nature of these cross-bridges is discussed: it is suggested that alpha-actinin possesses the characteristics for the cross-bridging protein and that vesiculation might be explained by the displacement of alpha-actinin by tropomyosin.

Animals↗
PubMed1974

A rapid method for the preparation of microvilli from rabbit kidney.

A simple method for the isolation of microvilli from kidney brush border is described. The method depends on the preferential aggregation of other subcellular structures by bivalent metal ions. MgCl(2) is added to a homogenate of cortical tissue prepared from frozen rabbit kidneys. Aggregated material is removed by a low-speed centrifugation and the supernatant centrifuged at 15000g to yield a pellet enriched in microvilli. This is resuspended and given a second treatment with Mg(2+). The purified preparation is obtained after four short differential centrifugations. The six brush-border enzymes that were monitored were enriched 11-17-fold compared with the original homogenate and were obtained in about 10% yield. Marker enzymes for other subcellular components showed the preparation to be essentially free of mitochondria and to be less contaminated with endoplasmic reticulum and baso-lateral plasma membranes than are conventional brush-border preparations. The main contamination was of lysosomal origin, about half of which was attributable to adsorbed acid hydrolases rather than to intact lysosomes. The aggregated components in the low-speed pellet bound less Mg(2+) than did the microvillus fraction. A possible mechanism for the role of Mg(2+) is discussed.

Animals↗
PubMedDate not supplied

An improved method for the preparation of human placental syncytiotrophoblast microvilli.

A simple procedure is described for the further purification of placental microvillus preparations. Based on previously published methods for the isolation of microvilli from other tissues, it depends on the preferential aggregation of containing structures by Mg2+. In the purified microvillus preparation, the two placental microvillar marker enzymes, alkaline phosphatase and 5'-nucleotidase, were enriched 24-fold and were obtained in 5 per cent yield. Five other microvilla enzymes were also further enriched by the Mg2+-treatment. Marker enzymes for other subcellular components showed that this treatment completely removed contamination by mitochondria and endoplasmic reticulum and contamination by lysosomes was decreased three-fold. (Na+ + K+)-activated ATPase was depleted by the Mg2+-treatment as was beta2-microglobulin.

5'-Nucleotidase↗
PubMedDate not supplied

Immunoelectrophoretic evidence that the human syncytiotrophoblast transferrin receptor is identical to a major plasma membrane antigen present throughout pregnancy.

Crossed immunoelectrophoresis of solubilized syncytiotrophoblast microvillar proteins has been performed through intermediate gels, containing a monospecific antiserum to transferrin, into gels containing an antiserum raised to purified microvilli from term placentae. This technique resolved the microvillar transferrin-receptor complex into its component proteins. Crossed hydrophobic interaction immunoelectrophoresis confirmed that the transferrin receptor is an amphiphilic integral membrane protein. It is also a major antigen of the microvillar membrane and is present throughout pregnancy.

Antigens↗
PubMedDate not supplied

Kidney microvillus peptidases--are they transmembrane proteins?

The membrane of kidney microvilli is richly endowed with peptidases. Present information is that there are at least eight examples located in this membrane. Three of the group are known to be among the major proteins that can be identified by dodecyl sulphate electrophoresis of the purified microvillus fraction. These three peptidases, aminopeptidase M, serine peptidase (dipeptidyl peptidase IV) and neutral endopeptidase can be labelled by lactoperoxidase iodination from either the luminal or the inner surfaces of the membrane, a result consistent with the view that the polypeptide chains span the microvillus membrane. The serine peptidase has been purified by two methods, permitting a comparison of the detergent-released and proteinase-released forms. The two forms differ in the presence and absence of the hydrophobic anchor that secures the enzyme to the membrane. Preliminary studies support the view that this hydrophobic domain is relatively small and that it includes the N-terminal region of the polypeptide chain.

Alanine↗