Multiple leucine aminopeptidases in the oocysts of Eimeria tenella and their changes during sporulation

• 1.1. There was little neutral protease activity but high levels of leucine aminopeptidases (LAP) in the oocysts of Eimeria tenella.
• 2.2. By electrophoretic analysis, there were three apparent LAP isozymes I, II and III in unsporulated oocysts.
• 3.3. They all diminished with the simultaneous emergence of a new, fast-moving isozyme V during late phase of sporulation.
• 4.4. The enzyme V was unlikely to have resulted from de novo protein synthesis and was predominantly in the cytoplasm surrounding the sporocysts.
• 5.5. It differed from the other isozymes by a slightly higher pH optimum, more dependence on Mn²⁺ or Mg²⁺ in the assay and higher susceptibility to chelating agents.
• 6.6. The possible biological function of these isozymes remain unknown. Since they were not found in sporozoites or merozoites of E. tenella, they may be needed only for sporulation and, possibly, excystation.

     

Characterization of proteins from mitochondrial ribosomes of Locusta migratoria by three different two-dimensional gel electrophoretic methods and comparison with cytoplasmic ribosomal proteins

• 1.1. Proteins were isolated from subunits of mitochondrial and cytoplasmic ribosomes of Locusta migratoria and were analyzed by means of two-dimensional gel electrophoreses using three different electrophoresis systems.
• 2.2. Using the system of Czempiel et al. (1976) proteins from whole locust mitochondrial ribosomes (combined subunits) were separated into 72 spots; proteins from the large and small subunits resulted in 48 and 29 spots respectively.
• 3.3. The mol. wt distribution of mitochondrial ribosome proteins was estimated by using the electrophoresis system of O'Farrell (1975). These mol. wts are in the range of 11,000–56,000, the average mol. wt is about 29,500. Assuming one copy of protein per ribosome this gives a total mol. wt for the protein part of mitochondrial ribosomes of ca. 2.1 x 10⁶.
• 4.4. Parallel separation of cytoplasmic and mitochondrial ribosome proteins was achieved using the system of Geyl et al. (1981). Cytoplasmic ribosome proteins produced 65 spots and revealed a more alkaline character than mitochondrial ribosome proteins.

     

The breakdown of Tetrahymena ribosomes by lysosomal enzymes: Inhibition by cytosol

• 1.1. Extracts from Tetrahymena lysosomes contained acid RNase and proteinase. At pH 7.4 there was appreciable proteinase activity which was inhibited by a heat-stable protein present in cell sap.
• 2.2. Lysosomal enzymes rapidly converted 80S ribosomes to subunits at pH 7.4. Hydrolysis of ribosomal RNA was very slow at pH 7.4 but rapid at pH 5.0.
• 3.3. These reactions were inhibited by proteinase inhibitors and by cell sap, but the latter was relatively ineffective at pH 5.0.
• 4.4. It seems unlikely that ribosome breakdown in vivo is initiated by the release of lysosomal enzymes into the cytosol.

     

Non-enzymic binding of phenylalanyl transfer ribonucleic acid to ribosomes and ribosomal subunits from rat liver

• 1.1. (³H)phe-(³²P)tRNA was used to compare the binding of phe-tRNA to 80 S ribosomes and 40 S subunits from rat liver in circumstances where deacylation of bound tRNA could be measured.
• 2.2. About twice as much phe-tRNA was bound to the 80 S ribosome as to the 40 S subunit.
• 3.3. Deacylation of phe-tRNA bound to the 80 S ribosome did not occur but about 20 % of the phetRNA bound to the 40 S subunit was lost by deacylation.

     

Purification of a proteinase and proteinase inhibitor from Tetrahymen a pyriformis

• 1.1. A cysteine proteinase and cysteine proteinase inhibitor have been purified from Tetrahymena.
• 2.2. The proteinase was purified by ammonium sulphate fractionation, gel filtration, ion exchange chromatography and affinity chromatography, and appeared homogeneous by gel filtration and electrophoresis (mol. wt approx 28,000). It hydrolysed BAPNA, degraded azocasein, and converted 80S ribosomes to subunits. Thiol reagents inhibited these activities.
• 3.3. The inhibitor was purified by heat treatment, ammonium sulphate fractionation and ion exchange chromatography, and appeared homogeneous by gel filtration and electrophoresis (mol. wt approx 12.500). The inhibitor was heat stable and it inhibited papain, as well as the Tetrahymena proteinase.

     

Amino acid composition and the protein form of procarboxypeptidase a purified from the pancreas of the sei whale Balaenoptera bolealis

• 1.1. Procarboxypeptidase (W-PCPA) was purified from the pancreas of the sei whale Balaenoptera bolealis.
• 2.2. W-PCPA was obtained as a homogeneous protein in polyacylamide gel disc electrophoresis.
• 3.3. W-PCPA has a molecular weight of 75,000.
• 4.4. Amino acid composition of W-PCPA was compared with that of bovine procarboxypeptidase as A S5 (PCPA-S5).
• 5.5. W-PCPA may be two subunits, and the aggregate form may resemble PCPA-S5.

     

Calcutta-1 LDH: Kinetic and thermodynamic properties of an electrophoretic variant of human LDH

• 1.1. Kinetic constants determined for the purified heterozygous variant LD₁ were closely similar to those of normal human LD₁.
• 2.2. Calcutta-1 homozygote LDH differed from normal LDH in K_m NADH and in Arrhenius activation energy.
• 3.3. The normal B subunits confer stability on the mutant subunits in the heterotetramers of Calcutta-1 LD₁.

     

S-Adenosyl-l-methionine decarboxylase activities in Mytilus edulis

• 1.1. The activities of S-adenosylmethionine decarboxylase (EC 4.1.1.50) were measured in cell extracts of mantle, hepatopancreas and foot from Mytilus edulis.
• 2.2. The apparent molecular weights of the enzymes estimated by gel filtration chromatography were 65,000 ± 10,000.
• 3.3. The enzymes do not require bivalent cations for catalysis and show optimum pH between 7.0–8.0 in phosphate buffer.
• 4.4. The hepatopancreas enzyme shows different behavior to the other two enzymes against temperature and its activity is strongly inhibited by NH₄⁺.
• 5.5. The apparent K_ms for S-adenosylmethionine were found to be 300, 200 and 250 μM for the hepatopancreas, mantle and foot enzymes, respectively.

     

Regulation of polyamine synthesis in chicken liver

• 1.1. Insulin injected into fasting chickens promotes a marked increase of liver S-adenosylmethionine decarboxylase, with maximum activity 6 hr after administration.
• 2.2. The apparent half-life of the enzyme is 22 min.
• 3.3. Ornithine and putrescine in vivo give rise only to small modifications in enzyme activity; spermidine and spermine induce the enzyme.
• 4.4. Putrescine activates the decarboxylase in vitro. Spermidine and spermine, however, appear to be inhibitors.
• 5.5. Insulin promotes a decrease of arginine, ornithine and all other amino acids, except for proline which increases. Putrescine is enhanced, spermidine slightly decreased and spermine unchanged.
• 6.6. Levels of polyamines are significantly altered by administration of methionine which promotes synthesis of spermidine, and by spermine which blocks transformation of putrescine into spermidine.

     

Fatty acid synthetase from pigeon red blood cells

• 1.1. Fatty acid synthetase has been purified 200-fold from pigeon erythrocytes.
• 2.2. The enzyme gave 2 major staining bands on disc gel electrophoresis corresponding to the complex and dissociated forms of the enzyme.
• 3.3. Sucrose density gradient centrifugation of the enzyme showed only one sedimenting peak and high performance liquid chromatography also showed only 1 major light absorbing peak.
• 4.4. The molecular weight of the enzyme was estimated to be 300,000–330,000 and the enzyme is comprised of 2 subunits of similar molecular weights.
• 5.5. The red blood cell fatty acid synthetase was found to be immunochemically nonidentical with the liver fatty acid synthetase.

     

Partial characterization and subunit analysis of major phosphoproteins of egg yolk in the fish,Oryzias latipes

• 1.1. Molecular weight estimation and subunit analysis of four yolk phosphoproteins (PP1-PP4) in medaka (Oryzias latipes) eggs were performed.
• 2.2. PP1 (M_r ≈ 210,000) and PP2 (M_r ≈ 180,000) were found to be heterodimers composed of subunits of 113,000 and 94,000 and subunits of 84,000 and 72,000, respectively.
• 3.3. PP3 and PP4 [phosvitins of medaka (Murakami et al., 1990, Devl. Growth Differ.32, 619–627)], were monomeric phosphoproteins having mol. wts of about 40,000 and about 20,000, respectively.
• 4.4. Lipid composition of the mixture of PP1 and PP2, vitellogenin and yolk were found to be almost the same. PP1 and PP2 are probably lipovitellins of medaka.

     

Ostrich fructose-1,6-bisphosphatase: Kinetic characterization of the liver isoenzyme

• 1.1. Purified ostrich (Struthio camelus) liver fructose-1,6-bisphosphatase exhibited an absolute requirement for Mg²⁺.
• 2.2. The enzyme catalyzed the hydrolysis of fructose-1,6-bisphosphate, sedoheptulose-l,7-bisphosphate and ribulose-l,5-bisphosphate.
• 3.3. S_0.5 for substrate was 1.4 μM.
• 4.4. AMP was a potent non-competitive inhibitor with respect to substrate (K_i of 25 μM).
• 5.5. Fructose-2,6-bisphosphate was a potent competitive inhibitor of the enzyme (K_i of 4.8 μM).

     

The carotenoids of eggs of wild and farmed cod (Gadus morhua)

• 1.1. Eggs of wild cod, and of farmed cod fed (a) a diet supplemented with astaxanthin and (b) a diet supplemented with both astaxanthin and canthaxanthin, were analysed with respect to carotenoids.
• 2.2. The total carotenoid contents in eggs were 0.7 ppm for wild cod and 0.5 ppm for farmed cod.
• 3.3. Cod, having white flesh, deposit ketocarotenoids in the eggs, preferably astaxanthin.
• 4.4. Canthaxanthin can replace astaxanthin in the eggs, but astaxanthin appears to be deposited preferentially when both carotenoids are present in the diet.
• 5.5. The isomer distribution of (3S, 3′S):(3R, 3′S, meso):(3R, 3′R) astaxanthin in the eggs reflected the isomer composition of the diet.
• 6.6. Echinenone, 4′-hydroxyechinenone, adonixanthin and zeaxanthin encountered in cod eggs may represent reductive metabolites of canthaxanthin and astaxanthin.

     

Subunits of Potamilla leptochaeta chlorocruorin

The dissociation of the extracellular chlorocruorin of Potamilla leptochaeta was investigated by polyacrylamide gel electrophoresis (PAGE) in the presence of sodium dodecyl sulfate (SDS).

• 1.1. The unreduced chlorocruorin dissociated into three subunits with molecular masses of 15,000 (1), 30,000 (2) and 65,000 (3); reduced chlorocruorin provided one broad band of molecular mass 14,000 ± 2000.
• 2.2. Reelectrophoresis of each of the three subunits in the presence of 2-mercaptoethanol provided one broad band having a molecular mass of about 13,000 ± 2000.
• 3.3. It is proposed that Potamilla chlorocruorin consists of interchain disulfide-bonded dimers and tetramers of polypeptide chains of ca. 14,000 ± 2000.
• 4.4. Comparison of the subunit relationships observed in annelid extracellular hemoglobins with those seen in chlorocruorins suggests that there are differences between the two molecules, insofar as aggregation of the smallest subunit is concerned: although it associates into dimers and trimers in hemoglobins, it forms dimers and tetramers in chlorocruorins.

     

An octopamine-sensitive adenylate cyclase in the central nervous system of Limulus polyphemus

• 1.1. Adenylate cyclase (E.C. 4.6.1.1) was assayed and shown to be present in the protocerebrum and circumesophageal ring of Limulus polyphemus.
• 2.2. The enzyme activity from both tissues is stimulated by fluoride and guanylnucleotides.
• 3.3. The circumesophageal ring, but not the protocerebrum, is responsive to octopamine.
• 4.4. Octopamine stimulation of the adenylate cyclase is reversed by phentolamine and dopamine.

     

A carotenoid-pheophytin-protein complex from the digestive juice of silkworm (Bombyx mori L.)

• 1.1. A soluble carotenoid-pheophytin-protein complex was purified from the digestive juice of fifth instar silkworm larvae raised on mulberry leaves.
• 2.2. The pigment-protein complex showed absorption maxima at 276, 429, 453, 481 and 670 nm. Major pigment components were identified as α-carotene, pheophytin a and b.
• 3.3. This complex has an acidic protein component having an isoelectric point of 4.6. The molecular weight was estimated to be 68,000 with four identical subunits.

     

Metabolism of ingested indole compounds in the gut of three species of heteroptera

• 1.1. All kinds of indole compounds used for the experiment were more or less metabolized in the gut of Dolycoris baccarum, Eurydema rugosum and Elasmostethus humeralis.
• 2.2. The metabolic pattern of the bugs was classified into four types (I–IV) for several indole compounds in the same way as for IAA.
• 3.3. The IAA metabolites in the excreta of the three species were probably the high-molecular compound combining with such substances as amino acids, sugars or proteins to some position of indole nucleus.
• 4.4. The crude excreta of E. humeralis fed with each of several indole compounds had a significant auxin activity.
• 5.5. Most of the metabolites of indole-3-acetaldehyde in the excreta of E. humeralis had also a significant auxin activity.
• 6.6. The significance of auxin metabolism in the gut of the bugs and the difference of auxin metabolism between aphids and bugs are discussed.

     

The subunit structure of Daphnia pulex hemoglobin

• 1.1. The extracellular hemoglobin of Daphnia pulex has an apparent molecular weight of 430,000–470,000 by gel chromatography and an S_20,w = 16.9 at pH 7.0.
• 2.2. Purified hemoglobin contains one heme per 18,000–20,000 g protein. The polypeptide chains are heterogeneous with mol. wts between 31,000–37,000. Some high mol. wt (M_r = 53,000–86,000) material is also present.
• 3.3. The hemoglobin dissociates at pH 10.5 in EDTA into 3S material which can be digested with subtilisin into 16,000 mol wt heme-containing polypeptide chains.
• 4.4. The amino acid composition of the intact hemoglobin is identical to that of the heme-containing fragments generated by proteolytic digestion of the 3S material.
• 5.5. These results are consistent with the hypothesis that D. pulex hemoglobin is composed of subunits containig two heme groups per 35,000 mol. wt polypeptide chain.

     

A New Tool to Reveal Bacterial Signaling Mechanisms in Antibiotic Treatment and Resistance

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Highlights

• •Quantitative (phospho)proteome analysis of antibiotic treatment in E. coli.
• •Largest bacterial phosphorylation catalogue.
• •Specific phosphorylation motifs changes during resistance development.
• •Phosphorylation mediated signaling could be a potential target for drug design.

     

Characterization of a blue astaxanthin protein from the carapace of the crayfish Astacus leptodactylus

• 1.1. A blue carotenoprotein (λ_max = 634 nm) containing astaxanthin as prosthetic group, was extracted and purified from the carapace of the crayfish Astacus leptodactylus.
• 2.2. The blue carotenoprotein contained (3S,3′S)-astaxanthin, (3R,3′S, meso)-astaxanthin and (3R,3′R)-astaxanthin in relative ratio 38:41:21.
• 3.3. The blue carotenoprotein had an approximate mol. wt of 440,000 (gel filtration) and 437,000 (gradient gel electrophoresis).
• 4.4. Sodium dodecyl sulphate polyacrylamide gel electrophoresis indicated the presence of two polypeptides of 19,600 and 18,600 daltons, with different mobility in polyacrylamide gel electrophoresis in the presence of 6 M urea.
• 5.5. At low ionic strength and in the presence of denaturing agents such as SDS, urea, extreme pH and heat, the blue complex showed a greater stability than most of the carotenoproteins studied to date.