No description available.

No description available.

Plants within the Caryophyllaceae, and certain other families, produce cyclic peptides (CPs) which generally consist of 5–12 proteinogenic amino acids. Until recently, very little was known about the biosynthesis of CPs in the Caryophyllaceae. Recently, in the Covello lab, two enzymes in Saponaria vaccaria were found to be involved in the processing of ribosome-derived linear precursors, giving rise to cyclic peptides. Thus, oligopeptidase 1 (OLP1) and peptide cyclase 1 (PCY1) are involved in the biosynthesis of segetalin A (a six-membered CP) from a 32 amino acid linear peptide precursor called presegetalin A1. PCY1 carries out the unusual cyclization reaction to form mature segetalin A from a linear intermediate. The purified recombinant PCY1, the first cloned plant enzyme whose function is peptide cyclization, was identified as a homologue of a prolyl-oligopeptidase from the S9 serine protease family. In principle, PCY1 performs an intra-molecular transpeptidation reaction to produce a CP. A homology-based structural model of PCY1 suggests that it has two domains, a catalytic α/β hydrolase domain and an unusual β- propeller domain. In an effort to define the substrate specificity of PCY1, a wide variety of synthetic peptide precursors were tested in assays and the results are discussed.

No description available.

No description available.

No description available.

No description available.

Heparin is a highly sulphated glycosaminoglycan. Approximately 70% of the polymer structure is represented by the disaccharide repeat unit (IdA-2S→GlcNS-6S). Variations with respect to the degree of sulphation, acetylation of the amino group and configuration of the uronic acid introduces extensive microheterogeneity within the polymer primary sequence. Heparin is not just a blood anticoagulant but has a wide ranging capability to interact with inorganic ions, proteins and drugs. Such interactions have been increasingly studied with specific reference to structure-function relationships. Heparin was subjected to a variety of chemical modifications including de-N-sulphation, de-N/O-sulphation, N-acetylation, N-propionylation, N-sulphation and carboxyl reduction. Partially modified polymers were synthesised using either less stringent solvolytic conditions or hydrochloric acid. The modified polymers were characterised using high-resolution ^1 3 C-NMR spectroscopy, Ir spectroscopy and acid-base titrations. Prior analysis of native heparin provided a set of reference spectra and titration profiles. Specific ion replacement and polarimetry were employed to study the interactions between heparin and cations. Polarimetry was further utilised to investigate the effect of polymer modification on the interaction with calcium and copper (II) ions. A study was conducted on the ability of the modified polymers to potentiate antithrombin inhibition of the cleavage of synthetic substrates by Factor Xa and thrombin. In addition, the influence of the modified heparin on capillary vessel growth in the chick CAM was investigated. Heparin and the chemically modified heparins were subjected to the bacterial enzymes heparinase II and heparinase III. Analysis of the degradation products was conducted using molecular exclusion chromatography and HPLC. The substrate specificity of the two lyases was assessed together with the potential of employing the enzymes, in conjunction with HPLC, to characterise heparins from various sources. The major conclusion are: 1) The interaction of calcium and copper (II) ions with modified polymers are fundamentally different with the amino group playing an important role in the copper (II) interaction. 2) Polysaccharide-catalysed inhibition of thrombin by antithrombin is more pronounced than for Factor AXa. The polysaccharides appear to play a subsidiary role through the formation of a simple electrostatic interaction with thrombin and antithrombin, thus bringing the protease and inhibitor together. 3) Heparinase II is active against a wide range of heparin-like polymers producing a variety of disaccharides and tetrasaccharides. Two specific glycosidic linkages are, however, resistant to the action of the enzyme. 4) Heparinase III catalyses the degradation of heparin-like polymers that are N-sulphated or N-acetylated but is inactive in those parts of the molecule in which a 2-0-sulphated uronic acid is present. 5) Enzyme-catalysed degradation followed by HPLC analysis is a viable method of characterising the disaccaride composition of heparins and heparin-like polymers.

The <i>de novo</i> formation of long-chain fatty acids and the reduction of long-chain fatty acids to the corresponding fatty alcohol were studied in the calanoid copepod <i>Calanus finmarchicus (Calanus</i>) and the fresh-water fish, <i>Trichogaster trichopterus</i> (the gourami). <i>De novo</i> biosynthesis of fatty acids by the enzyme fatty acid synthase (FAS) was localised in the 6.3 x 10⁶g-min supernatant ('cytosolic' fraction) of homogenates of whole <i>Calanus</i> and was characteristic of a Type 1 multienzyme complex. The formation of fatty acids required the presence of acetyl-CoA, malonyl-CoA and was specific for NADPH as the reductant. The main products of the reaction were the saturated long-chain fatty acids stearic acid and palmitic acid, accounting for 54% and 25% of the total fatty acid product, respectively. The alcohol-forming enzyme, NADPH-fatty acyl-CoA oxidoreductase, from both <i>Calanus</i> the gourami roe was membrane-associated. Differential ultracentrifugation showed that a 6.3 x 10⁶g-min pellet (the 'particulate' fraction) was most active in the formation of long-chain fatty acohols. The specific activity of the enzyme in the gourami roe particulate fraction was over 100-fold higher than that in the <i>Calanus</i> particulate fraction. Both systems required the presence of an acyl-CoA generating system in the form of exogenous CoASH, ATP and Mg⁺⁺. The reaction was specific for NADPH as the reductant in the case of the gourami roe system. However, NADH substituted for NADPH in the <i>Calanus</i> system. Palmitic acid, in the presence of an acyl-CoA generating system, was the most effective substrate with the 18:0, 18:1 and 22:1 substrates giving progressively lower activities.

Previous investigations of two direct methyltransferase systems in winter wheat have shown these systems to be operating at different times during the germination period and have suggested interconversions between these systems. S-Methyl-L-methionine, a methyl donor involved in these systems, could be transferring methyl groups to nucleic acid bases, histones, or pectic substances.In this study the translocation of labeled methyl groups from S-methyl-L-methionine-i4CH3 into nucleic acids, histones, or pectic substances was examined. Absorption of S-methyl-L-methionine-14CH3 was established and uptake of the labeled methyl group was indicated in the seed parts for 1, 2, and 3 days germination, and in the root and shoot for 3 days germination. Autoradiographic analysis of labeled tissue suggests translocation of the methyl group to meristematic regions and to the walls of the elongating cells in the region of cell elongation in root tissue.