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Biomedical subjects

S Clarke

Publications and source records attributed to S Clarke.

At least 379 records · Page 21Linked to original sources
PubMed1983

Protection against pneumococcal infection in mice conferred by phosphocholine-binding antibodies: specificity of the phosphocholine binding and relation to several types.

Some anti-phosphocholine antibodies protect mice against challenge with certain, but not all, pneumococcal types. We found that both their isotype and reactivity with the cell wall C-polysaccharide of encapsulated pneumococci, as measured by immunodiffusion, were important in predicting the protective activity of anti-phosphocholine antibodies. We propose that the specificity of the protective antibodies includes the backbone of the phosphocholine-containing structure.

Amino Acid Sequence↗
PubMed1983

Acquisition of expressive signing by autistic children: an evaluation of the relative effects of simultaneous communication and sign-alone training.

Two methods of training autistic children to use manual signs were compared. Two children, one mute and one capable of some verbal imitation, were taught to use signs as expressive labels for pictures of objects. Using an alternating treatments design, speed of sign acquisition was compared across two training conditions in which signs were presented either accompanied by, or without, the corresponding verbal label. In both conditions, the training procedure incorporated reinforcement, modeling, prompting, fading, and stimulus rotation. The efficacy of training in both treatment conditions was demonstrated by the use of a multiple baseline control across signs, but no clear differences in acquisition speed across conditions were apparent. Posttests conducted to assess stimulus control of signing, and learning of verbal labels when these were present in training, showed that the behavior of the imitative, but not the mute, child was controlled by the verbal stimuli. The implications of the results both for understanding deficits characteristic of autistic children and for developing appropriate language training procedures are discussed.

Adolescent↗
PubMed1982

A study of neurodevelopmental findings in early adolescent delinquents.

Subtle deficits or delays in the development of the central nervous system in young children are associated with problems affecting learning, behavior, and social adjustment. Less is known about the impact of such neurodevelopmental dysfunction in adolescents. This study assesses the possible association between neurodevelopmental delays and juvenile delinquency. An adolescent neurodevelopment examination was devised and administered to 54 delinquents and 51 secondary school students. Six areas were assessed. There were no significant differences in the prevalence of minor neurologic signs (P = 0.37) or in gross motor function (P = 0.02) and temporal sequential organization (P = 0.04). The greatest differences were in visual processing (P = 0.0002) and auditory-language function (P = 0.0001). Eighteen percent of delinquents and 4% of the comparison group were deficient in two or more neurodevelopmental areas. Nine percent of the delinquents and none of the comparison group had three or more dysfunctions. A neurodevelopmental examination may be a useful diagnostic tool for identifying endogenous factors in behavior, learning, and adjustment problems of this age group. Such findings have implications for the formulation of individualized management strategies.

Adolescent↗
PubMed1982

Methylation at D-aspartyl residues in erythrocytes: possible step in the repair of aged membrane proteins.

Reversibly methylated aspartyl residues in human erythrocyte membrane proteins are shown to be in the "unnatural" D configuration. This is demonstrated by treatment of proteolytically derived aspartic acid beta-[3H]methyl ester with L- and D-amino-acid oxidases and by the resolution of diastereomeric L-leucyl dipeptides containing either L- or D-aspartic acid beta-methyl ester by ion-exchange chromatography. Based on this observation, we propose a novel role for eukaryotic protein carboxyl methyltransferases (protein O-methyltransferase; S-adenosyl-L-methionine:protein O-methyltransferase, EC 2.1.1.24). We suggest that these widely distributed enzymes function to recognize aspartyl residues that have racemized spontaneously for a subsequent repair reaction. This repair function is postulated to couple ester hydrolysis with the restoration of the original L configuration of the aspartyl residue. It is possible that similar types of racemization repair processes may occur by reversible covalent modifications at other residues. Other possible functions for D-aspartic acid beta-methyl ester residues in proteins are considered.

Amino Acid Oxidoreductases↗
PubMed1981

Direct renaturation of the dodecyl sulfate complexes of proteins with Triton X-100.

A simple procedure is described for renaturing dodecyl sulfate-unfolded enzymes. The method involves the direct addition of a large molar excess of the non-ionic detergent Triton X-100 to protein-dodecyl sulfate complexes either in solution or as a band on a polyacrylamide gel. The cytoplasmic enzymes lactate dehydrogenase and malate dehydrogenase have been renatured by this protocol. On the other hand, no recovery of activity was found with the mitochondrial isoenzymes of malate dehydrogenase or the mitochondrial enzymes glutamate dehydrogenase or fumarase. Possible implications of the differences in the ability of cytosolic and mitochondrial enzymes to renature under these conditions are discussed in terms of their biosynthesis.

Cytosol↗
PubMed1981

Reversible methylation of cytoskeletal and membrane proteins in intact human erythrocytes.

Membrane protein methylation was studied in intact human erythrocytes. Erythrocytes were incubated with physiological concentrations of L-[methyl]3H]methionine and incorporated 35 pmol of methyl groups into membrane components/mg of membrane protein in a 2.5-hr incubation at 37 degrees C. At least 90% of the total groups (12,500 methyl groups/cell) were incorporated into polypeptides via linkages which were labile to 1 N NaOH. Major methylated membrane polypeptides were identified based on the comigration of radioactivity with Coomassie blue- and periodic acid-Schiff-staining species in pH 2.4 dodecyl sulfate gel electrophoresis, as well as by the distribution of radiolabel and protein following selective proteolysis and membrane extraction procedures. Methylated species identified in this way include the cytoskeletal polypeptides band 2.1 (ankyrin) and band 4.1, as well as the band 3 anion transport protein. Other methylated species include an intrinsic polypeptide comigrating with band 3 but insensitive to external chymotrypsin digestion, a glycoprotein showing variable migration in this gel system (40,000-55,000 daltons), an intrinsic polypeptide at about 30,000 daltons, and an extrinsic species of about 17,000 daltons. A small amount of radioactivity comigrated with the band 4.5 region. Bands 1, 2, 4.2, 5, 6, 7, and the major sialoglycoprotein were not methylated in this system. All of the methylated species exhibited turnover in vivo, and the time taken to reach 50% demethylation for each species ranged from less than 2 to 29 h.

Chymotrypsin↗
PubMed1981

Methylation of membrane proteins in human erythrocytes. Identification and characterization of polypeptides methylated in lysed cells.

An in vitro system was developed for studying protein methylation reactions in human red blood cells. Packed erythrocytes were lysed by freeze-thawing in the presence of S-adenosyl[methyl-3H]methionine. Specific incorporation of base-labile methyl groups into the band 3 anion transport protein and the major sialoglycoprotein (glycophorin, periodic acid-Schiff reagent-1) was demonstrated by dodecyl sulfate gel electrophoresis at pH 2.4, selective extractions with Triton X-100 and lithium diiodosalicylate, and protease sensitivity. Two other unidentified intrinsic membrane proteins with Mr = 96,000 and 23,500 were also methylated. Little radioactivity was incorporated into membrane proteins when membranes were incubated with S-adenosyl-L-[methyl-3H]methionine in the absence of cytosol. No evidence was obtained for incorporation of methyl label into extrinsic proteins including bands 1, 2, 2.1, 4, 5, 6, or in zone 4.5. Proteolytic digestion of intact cells and isolated membranes revealed that one site of methylation on the band 3 polypeptide may be at the inner surface of the membrane near the junction of the cytoplasmic domain and the membrane domain. The rates of hydrolysis of the incorporated methyl groups were characterized over a range of pH values. These rates were compared to those of methyl esterified amino acids and peptides, including aspartic acid beta-methyl ester which has been isolated from proteolytic digests of methylated erythrocyte membranes (Janson, C. A., and Clarke, S. (1980) J. Biol. Chem. 225, 11640-11643). We find that the rates of base-catalyzed hydrolysis of beta-methyl esters of aspartic acid and gamma-methyl esters of glutamic acid are highly sensitive to the presence of substituents on the alpha-carboxyl and alpha-amino groups. The rate of hydrolysis of the membrane-incorporated methyl groups are consistent with those of internal aspartic acid and glutamic acid methyl ester residues.

Chymotrypsin↗
PubMed1980

Identification of aspartic acid as a site of methylation in human erythrocyte membrane proteins.

Aspartic acid beta-[3H]methyl ester has been isolated from proteolytic digests of [3H]methylated human red blood cell membranes. The digestion product was identified by its co-elution with an ion exchange chromatography, gel filtration, and thin layer chromatography. The rate of hydrolysis of the methyl group of the isolated compound was determined at serveral pH values and was found to be identical with that of aspartic acid beta-methyl ester. This radioactive compound could be isolated from membranes prepared from broken cells incubated with S-adenosyl-L-[methyl-3H]methionine or from intact cells incubated with L-[methyl-3H]methionine. NO evidence was obtained for the presence of glutamic acid gamma-methyl ester in these digests. We suggest on the basis of these results that a major site of protein methylation in human red blood cell membrane proteins is at aspartyl residues.

Aspartic Acid↗
PubMed1980

In vitro methylation of bacterial chemotaxis proteins: characterization of protein methyltransferase activity in crude extracts of Salmonella typhimurium.

A specific in vitro assay was developed for the protein carboxyl methyltransferase that is involved in the chemotactic behavior of Salmonella typhimurium. This cytosolic enzyme catalyzes an S-adenosyl-L-methionine-dependent methyl esterification of glutamyl residues on a class of 60,000-dalton inner-membrane proteins. The activity was found to display a pH optimum of 6.5 and be sensitive to the concentration of salts in the assay medium. No detectable activity was found towards a variety of other proteins which serve as substrates for mammalian and other bacterial carboxyl methyltransferases. This assay was used to quantitate the methylation of the 60,000-dalton methyl-accepting proteins in response to chemoeffectors. Small but reproducible concentration-dependent changes in the initial rates of in vitro methylation were observed with chemotactic attractants and repellents. The specific methyltransferase activity was found to be absent in several mutants in flagellar synthesis (fla-), suggesting that the synthesis of this enzyme is coordinately regulated with that of flagellin and basal bodies. The hydrodynamic properties of the enzyme in crude extracts were determined by gel filtration and sucrose velocity gradient centrifugation, and a native molecular weight of 41,000 was calculated from these data.

Bacterial Proteins↗
PubMed1979

Membrane receptors for aspartate and serine in bacterial chemotaxis.

High affinity binding sites for serine and aspartate have been characterized in membranes from Salmonella typhimurium and Escherichia coli. Greater than 80% of these sites have been identified as chemotaxis receptors. Mutants lacking binding sites for these amino acids have been shown to have corresponding defects in taxis. The substrate specificity of each of the receptors in Salmonella is very high; most analogs of serine and aspartate do not bind to these receptor sites and do not affect chemotaxis. The transport of these amino acids is apparently not related to chemotaxis. At least 2500 serine receptors and 1200 aspartate receptors with dissociation constants of about 5 microM are present in the membrane fraction of logarithmically growing cells.

Aspartic Acid↗