Comparative Biochemistry and Physiology Part B: Comparative Biochemistry最新文献

A soluble protein in Rhesus monkey milk was isolated to apparent homogeneity by FPLC gel filtration, anion-exchange and reverse-phase chromatography. It is a major milk protein and is present at 2.5—3.0 mg/ml milk throughout lactation. It is only found in the whey fraction of milk; acid precipitation of casein does not result in any significant change in its concentration. A molecular weight (MW) of about 21.6 kDa was estimated from gel filtration and SDS gel electrophoresis and also calculated from its amino acid composition. The amino acid composition of this protein is similar to that of bovine β-lactoglobulin (β-Lg), but it is larger in size, possibly representing a family of primate β-Lgs.

A comprehensive study was performed on the brains of various vertebrate species showing different life energy potentials in order to find out if free radicals are important determinants of species-specific maximum life span. Brain superoxide dismutase, catalase, Se-dependent and independent GSH-peroxidases, GSH-reductase, and ascorbic acid showed significant inverse correlations with maximum longevity, whereas GSH, uric acid, GSSG/GSH, in vitro peroxidation (thiobarbituric acid test), and malondialdehyde (measured by HPLC), did not correlate with maximum life span. Superoxide dismutase, catalase, GSH-peroxidase, GSH and ascorbate results agree with those previously reported in various independent works using different animal species. GSSG/GSH, and true malondialdehyde (HPLC) results are reported for the first time in relation to maximum longevity. The results suggest that longevous species simultaneously show low antioxidant concentrations and low levels of in vivo free radical production (a low free radical turnover) in their tissues. The “free radical production hypothesis of aging” is proposed: a decrease in oxygen radical production per unit of O₂ consumption near critical DNA targets (mitochondria or nucleus) increases the maximum life span of extraordinarily long-lived species like birds, primates, and man. Free radical production near these DNA sites would be a main factor responsible for aging in all the species, in those following Pearl's (Rubner's) metabolic rule as well as in those not following it.

The expression of alleles encoding the enzymes lactate dehydrogenase (LDH; EC 1.1.1.27) and sorbitol dehydrogenase (SDH; EC 1.1.1.14) was investigated in the redtail rasbora (Rasbora borapetensis), the eyespot rasbora (R. dorsiocellata), and in their reciprocal hybrids, by acrylamide gel electrophoresis. The spatial and temporal expression of LDH and SDH isozymes in Rasbora is consistent with those reported for other teleosts. The asynchronous delay in appearance of isozymes of paternal and maternal LDH-C and SDH-A subunit composition is in keeping with observed effects of gene regulatory divergence where alleles of maternal origin interact more effectively with maternal cytoplasmic regulatory factors than do alleles of paternal origin.

The effect of seasonal acclimatization on the extent of prolactin (PRL) gene expression and on the content of this was studied in summer- and winter-carp (Cyprinus carpio) hormone pituitary glands. PRL content in the rostral pars distalis (RPD) was evaluated by immunocytochemistry using antibodies against a cross-linked synthetic peptide comprising the sequence of 15 amino acids which conform to the primary structure of carp PRL. To assess the level of PRL gene transcription, a 24-mer synthetic oligonucleotide probe whose sequence included nucleotides 2041–2064 located in exon V of the carp PRL gene, was used. Employing in situ hybridization assays, a high expression of PRL mRNA was observed in the RPD of summer-acclimatized carp. A negligible level of transcription was observed in tissue sections of pituitary glands from winter-acclimatized carp. Concurrently, immunodetection of the PRL-producing cells in the RPD revealed that the pituitary hormone level was significantly higher in the warm season-adapted carp.

Alanopine dehydrogenase, octopine dehydrogenase and lactate dehydrogenase activities were detected in the heart of C. concholepas. As determined by gel filtration on Sephadex G-100, the molecular weights (M_r) were 85,000, 42,000 and 52,000 for lactate dehydrogenase, alanopine dehydrogenase and octopine dehydrogenase, respectively. Substrate inhibition of the opine dehydrogenases was observed at high concentrations of both pyruvate and the corresponding amino acid (alanine or arginine). Moderate substrate inhibition of lactate dehydrogenase by pyruvate was observed; K_m values for lactate and NAD⁺ were unusually high. Alanoonine dehydrogenase was very specific for alanine and glycine was not a substrate for this enzyme. In addition to arginine, lysine was also a substrate for octopine dehydrogenase. Possible functions of the pyruvate reductases are discussed in connection with adaptation to anoxia and other regulatory processes in the heart of C. concholepas.

The land snail, Otala lactea, like many other organisms, depresses its metabolism in the face of environmental stress. Due to the extent of this depression, a decrease in the rate of protein synthesis is likely to be involved. In an attempt to quantitate changes in the rate of protein synthesis during metabolic depression in O. lactea we have determined polysome profiles from the hepatopancreas of awake and estivating snails. In both estivating and awake snails, the majority of ribosomes were present as monosomes and there was no evidence of qualitative changes with the state of the snail.

A simple and rapid purification procedure for hepatic peroxisomal multifunctional enzyme (Δ³,Δ²-enoyl-CoA isomerase/enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase) from clofibrate treated mice is described. The purification is achieved within two days using ion-exchange chromatography and an easily prepared affinity resin. The overall yield is 10% or more after a 100-fold enrichment from the cytosolic fraction of liver tissue. The native enzyme is a monomer with a molecular mass of 75 kDa. The protein is blocked in the N-terminus but internal amino acid sequences was obtained after proteolytic cleavage. Western blot analysis indicated proteolysis of multifunctional enzyme in different subcellular fractions derived from liver tissue. The hydratase activity of the enzyme is heat-labile and highly dependent on the concentration of Tris buffer or potassium chloride present. Optimal activity was found around 37°C and pH 7. The enzyme also shows dehydrogenase and isomerase activity.

A branched-chain acyl-CoA transferase activity which transfers coenzyme A from either 2-methylbutyryl or 2-methylvaleryl-CoA to succinate is present in the muscle mitochondria from the intestinal nematode, Ascaris suum. Its physiological function is discussed. This activity appears to differ from the previously described acetyl-CoA:propionate and propionyl-CoA:succinate acyl-CoA transferases on the basis of heat stability, substrate specificity and the requirement of a “factor” from boiled Ascaris mitochondria for optimal activity of only the branched-chain acyl-CoA transferase. The “factor” has been recovered from HPLC and some of its properties examined. It could not be replaced by a crude soluble fraction from rat liver mitochondria, or by adenine, guanine or inosine di- or triphosphates. Activity was lost upon ashing, but was not affected by treatment with either pepsin or chymotrypsin.

Lipid and phospholipid compositions of an endemic deep-water freshwater gammarid, belonging to the subphylum Crustacea, Acanthogammarus grewingkii was studied. Content of alkenylacyl, alkylacyl and diacyl forms in the main phospholipid classes (phosphatidylethanolamine and phosphatydilcholine) were established using reaction micro-thin-layer chromatography. The fatty acids compositions of total lipids, neutral, glyco- and phospholipid fractions were investigated by capillary gas chromatography-mass spectrometry. Seventy-nine fatty acids were identified: 26 saturated (iso-, anteiso- and cyclo-), 26 monoenoic, 7 dienoic, 14 trienoic and 16 tetra-, penta- and hexaenoic. A number of demospongic fatty acids, such as 5,9–25:2, 5,9,19–26:3, 5,9,17–26:3, 5,9,23–28:3 and 5,9,21–28:3 acids, were found.