DNA polymerases and DNA topoisomerases solubilized from nuclear matrices of regenerating rat livers

DNA topoisomerase activity together with the activities of DNA polymerase were detected in a form tightly associated with rat liver nuclear matrices. DNA polymerase activities were solubilized from the nuclear matrices of regenerating rat livers by sonic treatment followed by extraction of these activities with detergent and salt. The predominant activity was mainly α-polymerase as judged from the size determined by sucrose density gradient centrifugation. However, only β-polymerase activity was detected in the matrix of normal rat livers. DNA topoisomerase activity, detected in both regenerating and normal liver nuclear matrices, showed a molecular size of about 4 S in sucrose gradient, and was active in the presence of EDTA. These results suggest that this enzyme belongs to type I topoisomerase.     

An apparatus for simultaneous manual solid-phase synthesis of multiple peptide analogs

A new design of reaction vessel for simultaneous manual solid-phase synthesis of multiple peptide analogs is described. Simultaneous use of four of these vessels attached to a single rotary mixer has been successfully applied to synthesis of two sets of four decapeptide amide analogs. Efficient coupling was indicated by chemical determination at the end of each synthesis cycle and overall final yields of between 78 and 84% were obtained. The products obtained were of a high quality, as assessed by high-performance liquid chromatography and amino acid analysis. This system allows expeditious synthesis of multiple peptide analogs for structure-function studies with economical use of efficiently ventilated laboratory space.     

Complete amino acid sequence of human seminal plasma beta-inhibin. Prediction of post Gln-Arg cleavage as a maturation site

The complete sequence of a 94 amino acid human seminal plasma polypeptide exhibiting inhibin-like activity is presented. This molecule, called beta-inhibin, selectively and specifically suppresses the release of pituitary FSH in vivo as well as in vitro. It does not affect the secretion of LH. Such a novel acidic protein contains a very basic C-terminal segment which is easily cleaved by mild tryptic digestion. It is predicted that the FSH inhibiting activity may reside within this region of the molecule. This would imply a post Gln-Arg cleavage to release the basic C-terminal active moiety.     

The glycoprotein 71 of ecotropic Friend murine leukemia virus. Structure of the oligosaccharides linked to asparagine-12

The glycoprotein from Friend murine leukemia virus was digested with protease from Staphylococcus aureus V8. A glycopeptide comprising the N-terminal glycosylation site (Asn-12) was isolated from the mixture of fragments and analyzed by amino acid sequencing and methylation-capillary gas chromatography-mass spectrometry before and after treatment with sialidase from Vibrio cholerae. Asn-12 was thus found to be substituted by a family of partially sialylated, fucosylated, and intersected glycoprotein N-glycans of the hybrid type.     

Relationship of lipoamide dehydrogenases from Pseudomonas putida to other FAD-linked dehydrogenases

Pseudomonas putida produces two lipoamide dehydrogenases, LPD-glc and LPD-val. LPD-val is specifically required as the lipoamide dehydrogenase of branched-chain keto acid dehydrogenase and LPD-glc fulfills all other requirements for lipoamide dehydrogenase. Both proteins are dimers with one FAD per subunit. LPD-glc has an absorption maximum at 455 nm, but LPD-val has a maximum at 460 nm. Comparison of amino acid compositions revealed that LPD-glc was more closely related to Escherichia coli and pig heart lipoamide dehydrogenase than to LPD-val. LPD-val did not appear to be closely related to any of the proteins compared with the possible exception of mercuric reductase.     

Reduced maximum capacity of glycolysis in brown adipose tissue of genetically obese, diabetic (db/db) mice and its restoration following treatment with a thermogenic beta-adrenoceptor agonist

The maximal activities of the key glycolytic enzymes hexokinase and 6-phosphofructokinase, were reduced in brown adipose tissue in db/db mice compared to their lean littermates. Treatment of db/db mice with the thermogenic beta-adrenoceptor agonist, BRL 26830, restored normoglycaemia. The only significant increase in activity of hexokinase and 6-phosphofructokinase in the BRL 26830-treated db/db mice occurred in brown adipose tissue where the total tissue activity increased 10- and 11-fold respectively. These changes together with increased 2-deoxyglucose uptake in vivo suggest that brown adipose tissue can play a quantitatively important role in the removal of glucose from the blood.     

Monovalent carboxylic ionophores inhibit transport of carbamoyl-phosphate synthetase I into mitochondria in Reuber hepatoma H-35 cells and cause accumulation of enzyme precursor

Transport of the precursor for carbamoyl-phosphate synthetase I into mitochondria in Reuber hepatoma H-35 cells was inhibited by adding monensin or nigericin to the culture medium at a concentration of 0.5 microM, and the enzyme precursor accumulated, mainly in the cytosolic fraction. Accumulated precursor was degraded slowly with a half-life of more than 16 min. Valinomycin, nonactin, A23187, X-537A (lasalocid), bromo-lasalocid, and carbonyl cyanide m-chlorophenylhydrazone did not exhibit these effects at concentrations at which they did not inhibit protein synthesis of the cells.     

Protein-free models for the proton-coupled reduction of haemoproteins

Reduction of the bis-pilocarpate-haemin complex at pH greater than or equal to 10 involves the simultaneous uptake of an electron by the Fe(III) ion and a proton by the pendant alkoxide group of an axial ligand. This provides a protein-free model for reactions such as the proton-coupled reduction of cytochromes which involve cooperative Coulombic interaction between two non-bonded sites.     

Conservation of interphase chromatin nonhistone antigens as components of metaphase chromosomes

The degree of conservation of HeLa interphase chromatin nonhistone antigens among the nonhistones of isolated metaphase chromosomes was determined with immunological procedures. Proteins were separated on SDS-polyacrylamide gels and electrophoretically transferred to diazophenylthioether (DPT)-paper, which was then overlaid with antiserum to chromatin from interphase nuclei. The bound antibodies were detected with 125I-labeled protein A. Alternatively, polyacrylamide gels were directly overlaid with antiserum and with 125I-protein A. Densitometry of autoradiograms and stained gels revealed the degree of conservation of nonhistone antigenic determinants from interphase to metaphase to be over 90% for chromatin.     

Metabolite fluxes across the inner membrane of plant mitochondria — inhibition by phthalonic acid

Transport and oxidation-reduction of citrate, 2-oxoglutarate and oxaloacetate by mitochondria isolated from thermogenic (Arum maculatum, Sauromatum guttatum spadices), green leaf (Pisum sativum) or etiolated (Phaseolus aureus, Helianthus tuberosus) plant tissues was found to be inhibited by phthalonic acid. No inhibition was found for NADH oxidation, glutamate, succinate or glycine transport and oxidation and malate transport. The much greater sensitivity of citrate oxidation to phthalonate inhibition compared with that of 2-oxoglutarate indicated that different carriers were involved, neither of which appeared to be rate-limiting for oxidation. Fluxes of oxaloacetate, and their sensitivity to phthalonate, indicated that this keto acid may use either the same carrier as 2-oxoglutarate or an oxaloacetate-specific carrier.Abbreviation PTA phthalonic acid