ABSTRACT
Poly-beta-leucines have been evaluated as catalysts for the Juliá-Colonna asymmetric epoxidation of enones; the beta 3-isomer was found to be an effective catalyst for the epoxidation of chalcone (70% ee) and some analogues.
Subject(s)
Epoxy Compounds/chemical synthesis , Leucine/analogs & derivatives , Leucine/chemistry , Oligopeptides/chemistry , Catalysis , Chalcone/analogs & derivatives , Ketones/chemistry , StereoisomerismABSTRACT
The abilities of five polyamino acids (Paa's) to catalyse the asymmetric epoxidation of enones 1-7 under three sets of reaction conditions were compared: polyneo-pentylglycine and polyleucine showed distinct advantages in most circumstances. All five polymers were adsorbed onto silica and from this further study, immobilised polyneo-pentylglycine (PLNSi) and polyleucine (PLLSi) were shown to be the catalysts of choice for the asymmetric epoxidation of less-reactive alpha,beta-unsaturated ketones.
Subject(s)
Amino Acids/chemistry , Catalysis , Chemistry/methods , Ketones/chemistry , Models, Chemical , Oxygen/metabolism , Peptides/chemistry , Silicon Dioxide/chemistryABSTRACT
When using proteases in direct reversal of their normal hydrolytic function, the equilibrium position is very important in limiting the attainable yield in equilibrium-controlled enzymic peptide synthesis. Analysis of the equilibrium position reveals a favourable shift towards the peptide product if starting materials are largely undissolved in the reaction medium and the product precipitates. This approach enabled us to obtain high peptide yields in thermolysin-catalysed reactions in high-density aqueous media with an equimolar supply of substrates. The easy scale-up (up to mol-scale) of this approach is demonstrated by two examples. Z-His-Phe-NH2 and Z-Asp-Phe-OMe, precursors for cyclo-[-His-Phe-] and the low-calorie sweetener Aspartame, respectively, were synthesized in preparative yields of 84-88%.
Subject(s)
Dipeptides/chemical synthesis , Thermolysin/chemistry , Catalysis , Chromatography, High Pressure LiquidABSTRACT
The non-proteinogenic amino acid L-ornithine (L-Orn) can be conveniently obtained by enzymatic hydrolysis of arginine (Arg) with arginase (EC 3.5.3.1). Arginase from calf liver (Vmax = 459 mumol/(min.mg), Km = 25.5 mM) is inhibited competitively by L-Orn (Ki = 480 mM). The enzyme was found to be completely enantioselective (E-value > 100) so that D,L-Arg can be split into D-Arg and L-Orn. Operational stability at 25 degrees C (deactivation rate constant kdeact = 3.8 x 10(-3) h-1; tau 1/2 = 182 h) is sufficient for use in a continuous process but is significantly smaller than temperature stability (kdeact = 4.1 x 10(-4) h-1; tau 1/2 = 1682 h); mechanical stress through stirring and unsteady Mn2+ supply owing to oxidation in the continuous process are believed to cause the difference. Addition of ascorbic acid stabilizes calf liver arginase at temperatures higher than 25 degrees C (at 60 degrees C, delta delta G not equal to = 2.9 kJ/mol).
Subject(s)
Arginase/metabolism , Arginine/metabolism , Liver/enzymology , Ornithine/metabolism , Animals , Arginase/antagonists & inhibitors , Arginase/chemistry , Arginine/chemistry , Cattle , Enzyme Activation , Enzyme Stability , Hydrogen-Ion Concentration , Kinetics , Molecular Structure , Ornithine/chemistry , Ornithine/pharmacology , Regression Analysis , Stereoisomerism , Substrate SpecificityABSTRACT
The method of measuring enzyme deactivation by monitoring necessary addition of fresh enzyme to keep a constant degree of conversion in a CSTR at constant [E] x tau, the product of concentration of active enzyme [E] and residence time tau, was successfully applied to acylase I from porcine kidney and Aspergillus oryzae fungus. Fungal enzyme was found to be more stable than kidney enzyme. Activation by both Co2+ and Zn2+ ions also yielded increased operational enzyme stability: Co2+ and Zn2+ are better stabilizers than activators. Mg2+ and Ca2+ are found to be neither activators nor stabilizers. Fungal acylase partially deactivated by exposition to a metal-free medium in the CSTR was reactivated by addition of Zn2+, demonstrating that loss of Zn2+ from the enzyme molecule is mainly responsible for deactivation in a continuous reactor.
Subject(s)
Amidohydrolases/chemistry , Amino Acids/isolation & purification , Amidohydrolases/metabolism , Amino Acids/chemistry , Animals , Aspergillus oryzae/enzymology , Cations, Divalent , Enzyme Activation , Kidney/enzymology , Protein Denaturation , Stereoisomerism , SwineABSTRACT
L-ornithine has growth potential for parenteral nutrition and as a component for biologically active peptides. A process for enzymatic conversion of L-arginine to L-ornithine with arginase has been developed and tested on a pilot scale. With activation of arginase by Mn2+ and stabilization by ascorbic acid, the enzyme is sufficiently active and stable for application in large-scale L-ornithine synthesis.
Subject(s)
Arginase/chemistry , Arginine/chemistry , Ornithine/chemical synthesis , Biotechnology , Enzyme Stability , Hydrogen-Ion Concentration , Hydrolysis , Kinetics , Ornithine/chemistry , TemperatureABSTRACT
Several amino acid derivatives with the negatively charged N alpha-protecting groups Maleyl (Mal) and Citraconyl (Cit) were synthesized and used in enzyme-catalyzed peptide synthesis. Compared to commonly used alpha-amino protecting groups in chemical peptide synthesis (Z, Fmoc, Boc, etc.), these charged protecting groups strongly increase both water solubility of different aromatic amino acid derivatives and activities of synthesis reactions. As a consequence of the solubilizing effect, we used these groups in the kinetically controlled peptide synthesis choosing kyotorphin (tyrosyl-arginine) as a model peptide. With Mal-Tyr-OEt as substrate and Arg-OEt as nucleophile, we succeeded in the alpha-chymotrypsin-catalyzed production of about 12 kg of Tyr-Arg (50.4% overall yield) in a 300 l batch experiment.
