Wilfred L.F. Armarego

Bio: Wilfred L.F. Armarego is an academic researcher from Australian National University. The author has contributed to research in topics: Dihydropteridine Reductase & Pterin. The author has an hindex of 15, co-authored 69 publications receiving 10992 citations. Previous affiliations of Wilfred L.F. Armarego include Kyoto Women's University.

Purification of laboratory chemicals

Abstract: Common physical techniques used in purification chemical methods used in purification purification of organic chemicals purification of inorganic and metal organic chemicals general methods for the purification of classes of compounds purification of natural products biochemicals and related products.

Isolation and nucleotide sequence of the hmp gene that encodes a haemoglobin-like protein in Escherichia coli K-12.

Abstract: In the course of an attempt to identify genes that encode Escherichia coli dihydropteridine reductase (DHPR) activities, a chromosomal DNA fragment that directs synthesis of two soluble polypeptides of Mr 44000 and 46000 was isolated. These proteins were partially purified and were identified by determination of their N-terminal amino acid sequences. The larger was serine hydroxymethyltransferase, encoded by the glyA gene, while the smaller was the previously described product of an unnamed gene closely linked to glyA, and transcribed in the opposite direction. Soluble extracts of E. coli cells that overproduced the 44 kDa protein had elevated DHPR activity, and were yellow in colour. Their visible absorption spectra were indicative of a CO-binding b-type haemoprotein that is high-spin in the reduced state. The sequence of the N-terminal 139 residues of the protein, deduced from the complete nucleotide sequence of the gene, had extensive homology to almost all of Vitreoscilla haemoglobin. We conclude that E. coli produces a soluble haemoglobin-like protein, the product of the hmp gene (for haemoprotein). Although the protein has DHPR activity, it is distinct from the previously purified E. coli DHPR.

Chapter 4 – Purification of Organic Chemicals

Abstract: Publisher Summary This chapter discusses the general principles, techniques, and methods of purification of organic chemicals. The chapter includes commercially available organic chemicals. Most organic liquids and a number of solids can readily be purified by fractional distillation, usually at atmospheric pressure. Sometimes, particularly with high boiling or sensitive liquids, or when in doubt about stability, distillation or fractionation under reduced pressure should be carried out. Rapid purification procedures are also focused in this chapter for commonly used solvents and reagents, which make them suitable for general use in synthetic chemistry. Also enlisted are the molecular weights (to the first decimal place), melting points and/or boiling points together with the respective densities and refractive indexes for liquids, and optical rotations when the compounds are chiral. The chapter includes commercially available organic chemicals. Most of the inorganic, metalorganic, organo- bismuth, boron, phosphorus, selenium, silicon and alkali metal compounds and metal ion salts of organic acids are considered.

Purification of Organic Chemicals

Abstract: This chapter discusses the general principles, techniques, and methods of purification of organic chemicals. The chapter includes commercially available organic chemicals. Most organic liquids and a number of solids can readily be purified by fractional distillation, usually at atmospheric pressure. Sometimes, particularly with high boiling or sensitive liquids, or when in doubt about stability, distillation or fractionation under reduced pressure should be carried out. Rapid purification procedures are also focused in this chapter for commonly used solvents and reagents, which make them suitable for general use in synthetic chemistry. Also enlisted are the molecular weights (to the first decimal place), melting points and/or boiling points together with the respective densities and refractive indexes for liquids, and optical rotations when the compounds are chiral. The chapter includes commercially available organic chemicals. Most of the inorganic, metalorganic, organo- bismuth, boron, phosphorus, selenium, silicon and alkali metal compounds and metal ion salts of organic acids are considered.

Chapter 4 – Purification of Organic Chemicals

Abstract: Publisher Summary This chapter discusses the general principles, techniques, and methods of purification of organic chemicals. The chapter includes commercially available organic chemicals. Most organic liquids and a number of solids can readily be purified by fractional distillation, usually at atmospheric pressure. Sometimes, particularly with high boiling or sensitive liquids, or when in doubt about stability, distillation or fractionation under reduced pressure should be carried out. Rapid purification procedures are also focused in this chapter for commonly used solvents and reagents, which make them suitable for general use in synthetic chemistry. Also enlisted are the molecular weights (to the first decimal place), melting points and/or boiling points together with the respective densities and refractive indexes for liquids, and optical rotations when the compounds are chiral. The chapter includes commercially available organic chemicals. Most of the inorganic, metalorganic, organo- bismuth, boron, phosphorus, selenium, silicon and alkali metal compounds and metal ion salts of organic acids are considered.

Geometric and Electronic Structure/Function Correlations in Non-Heme Iron Enzymes

Abstract: ion step follows the decarboxylation, which is consistent with the deuterium isotopic effects observed for thymine 7-hydroxylase which indicate that an irreversible step (or steps) occurs prior to the C-H bond breaking.395 It has also been shown for prolyl 4-hydroxylase that a substrate-derived radical is generated in the reaction, which is consistent with a rebound mechanism.437 It is important to point out that no oxygen intermediate (i.e., bridged superoxo or oxo-ferryl) has been observed for any R-KGdependent enzyme. This warrants future theoretical and experimental study. A detailed molecular mechanism has been proposed for IPNS based on spectroscopic and crystallographic studies.422 Resting IPNS/FeII is also 6C and thus relatively stable toward dioxygen. Substrate ACV binds directly to FeII IPNS through its thiolate group, providing an open coordination position at the FeII. O2 can then react to form an FeIII-superoxo intermediate. This intermediate is suggested422 to perform the first hydrogen-atom abstraction step and close the â-lactam ring, resulting in the formation of the first water molecule and generating an FeIVdO-II intermediate, which completes the second ringclosure process by hydrogen-atom abstraction forming a thiazolidine ring. Previously proposed mechanisms of ACCO involved direct binding of cosubstrate ascorbate to the iron before O2 as part of the oxygen activation process.438,439 The EPR and ESEEM studies of the NO complex of ACCO suggested a quite different molecular mechanism for ACCO.435 An FeIII-superoxo intermediate is proposed. Whether it is preceded by a 6C f 5C process with substrate binding is presently under study.440 This intermediate is thought to initiate a radical process by single hydrogen-atom abstraction or electron-coupled proton transfer (PT)ion or electron-coupled proton transfer (PT) from the bound amino group. The resulting substrate radical may undergo spontaneous conversion into products. The role of cosubstrate ascorbate is proposed to reduce the toxic peroxo byproduct to water. Alternatively, the two-electron reduction of FeIIIsuperoxo by the cosubstrate ascorbate could result in an FeIVdO-II intermediate which initiates the radical reaction.435 4. Rieske-Type Dioxygenases Biochemical Characterization. The Rieske ironsulfur center is a two iron-two sulfur cluster ([2Fe2S]) which has a 2His (on one iron), 2Cys (on the other iron) coordination environment, instead of the 4Cys present in plant ferredoxins. It plays a key role in the electron transport pathway in membranebound cytochrome complexes as well as in some dioxygenases.441 The latter are mainly comprised of two protein components: a reductase containing flavin and a ferredoxin [2Fe-2S], and a terminal oxygenase containing a Rieske [2Fe-2S] cluster and a non-heme iron active site.442 Except for the recently reported alkene monooxygenase that has a binuclear iron site in its terminal oxygenase,10 most of the Rieske-type oxygenases have a mononuclear iron site, which is believed to be the site of dioxygen activation and substrate oxygenation.442,443 The majority of the Rieske-type mononuclear non-heme oxygenases form a family of enzymes which are aromatic-ring-hydroxylating dioxygenases. These catalyze the regioand stereospecific cis-dihydroxylation of an aromatic ring using dioxygen and NAD(P)H (Table 1). Examples include benzene dioxygenase (BDO, EC 1.14.12.3),444 phthalate dioxygenase (PDO, EC 1.14.12.7),445 toluene dioxygenase (EC 1.14.12.11),446 and naphthalene 1,2-dioxygenase (NDO, EC 1.14.12.12),447 which initiate the aerobic degradation of aromatic compounds in the soil bacteria and are targets for bioengineering in bioremediation. This step is the first step in the pathway that ultimately leads to ring cleavage by the intraand extradiol dioxygenases (sections II.B.2 and II.C.1).443 Besides these bacterial dioxygenases, other Rieske-type mononuclear non-heme oxygenases include anthranilate 1,2-dioxygenase (EC 1.14.12.1),448 which deaminates and decarboxylates the substrate to produce catechol; chlorophenylacetate 3,4-dioxygenase (EC 1.14.2.13),449 which converts substrate to catechol with chloride elimination; and 4-methoxybenzoate O-demethylase (putidamonooxin),450 which catalyzes the conversion of 4-methoxybenzoic acid to 4-hydroxybenzoic acid and formaldehyde. The reductase component is usually a monomer (MW ) 12-15 kDa) and utilizes flavin to mediate ET from the two-electron donor NAD(P)H to the oneelectron acceptor [2Fe-2S] cluster and is specific to each terminal oxygenase; other electron donors do not support efficient oxygenation.442 The crystal structure of phthalate dioxygenase reductase is available.451 The terminal oxygenases are large protein aggregates (MW ) 150-200 kDa) containing either multiples of R subunits (BDO R2, PDO R4) or an equimolar combination of R and â subunits (toluene dioxygenase R2â2, NDO R3â3). The R subunits contain a Rieske [2Fe-2S] cluster and a catalytic non-heme FeII center. â subunits do not seem to be involved in the catalytic function (vide infra). Kinetics. Steady-state kinetic studies coupled with various rapid reaction studies of the partial reactions of PDO allowed Ballou et al. to propose a kinetic scheme (Scheme 15).443 On the basis of steady state 278 Chemical Reviews, 2000, Vol. 100, No. 1 Solomon et al.

Tetrahydrobiopterin biosynthesis, regeneration and functions.

Abstract: Tetrahydrobiopterin (BH % ) cofactor is essential for various processes, and is present in probably every cell or tissue of higher organisms. BH % is required for various enzyme activities, and for less defined functions at the cellular level. The pathway for the de noao biosynthesis of BH % from GTP involves GTP cyclohydrolase I, 6-pyruvoyl-tetrahydropterin synthase and sepiapterin reductase. Cofactor regeneration requires pterin-4a-carbinolamine dehydratase and dihydropteridine reductase. Based on gene cloning, recombinant expression, mutagenesis studies, structural analysis of crystals and NMR studies, reaction mechanisms for the biosynthetic and recycling enzymes were proposed. With regard to the regulation of cofactor biosynthesis, the major controlling point is GTP cyclohydrolase I, the expression of which may be under the control of cytokine induction. In the liver at least, activity is inhibited by BH % , but stimulated by phenylalanine through the GTP cyclohydrolase I feedback regulatory protein. The enzymes that depend on BH % are the phenylalanine, tyrosine and tryptophan hydroxylases, the latter