Overexpression, purification and characterization of two pyrimidine kinases involved in the biosynthesis of thiamin: 4-amino-5-hydroxymethyl-2-methylpyrimidine kinase and 4-amino-5-hydroxymethyl-2-methylpyrimidine phosphate kinase

The overexpression, purification and characterization of 4-amino-5-hydroxymethyl-2-methylpyrimidine kinase (HMP kinase) and 4-amino-5-hydroxymethyl-2-methylpyrimidine monophosphate kinase (HMP-P kinase) are described. Surprisingly HMP-P kinase also shows HMP kinase activity. These enzymes are useful reagents for the preparation of intermediates on the thiamin biosynthetic pathway.     

PtMe(iPr3P)2]+: a Pt(II) complex with an agostic interaction that undergoes C-H activation

The T-shaped Pt(II) complex [PtMe(iPr3P)2][1-H-closo-CB11Me11], which is stabilised by an agostic interaction, undergoes acid-catalysed intramolecular C-H activation in the presence of THF to afford cyclometallated [Pt(THF)(iPr3P)(iPr2PCHMeCH2)][1-H-closo-CB11Me11].     

Synthesis and structural characterisation of rhodium hydride complexes bearing N-heterocyclic carbene ligands

Addition of excesses of N-heterocyclic carbenes (NHCs) IEt₂Me₂, IⁱPr₂Me₂ or ICy (IEt₂Me₂ = 1,3-diethyl-4,5-dimethylimidazol-2-ylidene; IⁱPr₂Me₂ = 1,3-diisopropyl-4,5-dimethylimidazol-2-ylidene; ICy = 1,3-dicyclohexylimidazol-2-ylidene) to [HRh(PPh₃)₄] (1) affords an isomeric mixture of [HRh(NHC)(PPh₃)₂] (NHC = IEt₂Me₂ (cis-/trans-2), IⁱPr₂Me₂ (cis-/trans-3), ICy (cis-/trans-4) and [HRh(NHC)₂(PPh₃)] (IEt₂Me₂(cis-/trans-5), IⁱPr₂Me₂ (cis-/trans-6), ICy (cis-/trans-7)). Thermolysis of 1 with the aryl substituted NHC, 1,3-dimesityl-4,5-dihydroimidazol-2-ylidene (IMesH₂), affords the bridging hydrido phosphido dimer, [{(PPh₃)₂Rh}₂(μ-H)(μ-PPh₂)] (8), which is also the reaction product formed in the absence of carbene. When the rhodium precursor was changed from 1 to [HRh(CO)(PPh₃)₃] (9) and treated with either IMes (=1,3-dimesitylimidazol-2-ylidene) or ICy, the bis-NHC complexes trans-[HRh(CO)(IMes)₂] (10) and trans-[HRh(CO)(ICy)₂] (11) were formed. In contrast, the reaction of 9 with IⁱPr₂Me₂ gave [HRh(CO)(IⁱPr₂Me₂)₂] (cis-/trans-12) and the unusual unsymmetrical dimer, [(PPh₃)₂Rh(μ-CO)₂Rh(IⁱPr₂Me₂)₂] (13). The complexes trans-3, 8, 10 and 13 have been structurally characterised.     

Mononuclear and dinuclear complexes with a [Ru(tBu2PCH2CH2PtBu2)(CO)] core

Thermolysis of solid [Ru(d(t)bpe)(CO)2Cl2](2, d(t)bpe =(t)Bu2PCH2CH2P(t)Bu2) under vacuum affords the five-coordinate complex [Ru(d(t)bpe)(CO)Cl2] (4), which was shown by X-ray crystallography to contain a weak remote agostic interaction. In solution, 4 can be readily trapped by CO, CH3CN or water to give [Ru(d(t)bpe)(CO)(L)Cl2](L = CO, 2; L = CH3CN, 6; L = H2O, 7). Reaction of 4 with AgOTf/H2O yields the tris-aqua complex [Ru(d(t)bpe)(CO)(H2O)3](OTf)2 (8), which has been structurally characterised and probed in solution by pulsed-gradient spin echo (PGSE) NMR spectroscopy. The water ligands in 8 are labile and easily substituted to give [Ru(d(t)bpe)(CO)(NCCH3)3](OTf)2 (10) and [Ru(d(t)bpe)(CO)(DMSO)3](OTf)2 (11). In the presence of CO, the tris-aqua complex undergoes water-gas shift chemistry with formation of the cationic hydride species [Ru(d(t)bpe)(CO)3H](OTf) (12) and CO2. X-Ray crystal structures of complexes 2, 4, 6, 8 and 11-12 are reported along with those for [{Ru(d(t)bpe)(CO)}2(mu-Cl)2(mu-OTf)](OTf) (3), [{Ru(d(t)bpe)(CO)}2(mu-Cl)3][Ru(d(t)bpe)(CO)Cl3](5) and [Ru(d(t)bpe)(CO)(H2O)2(OTf)](OTf)(9).     

Synthesis, characterisation and optical spectroscopy of platinum(II) di-ynes and poly-ynes incorporating condensed aromatic spacers in the backbone

A series of protected and terminal dialkynes with extended pi-conjugation through a condensed aromatic linker unit in the backbone, 1,4-bis(trimethylsilylethynyl)naphthalene, 1,4-bis(ethynyl)naphthalene, 9,10-bis(trimethylsilylethynyl)anthracene, 9,10-bis(ethynyl)anthracene, have been synthesized and characterized spectroscopically. The solid-state structures of and have been confirmed by single crystal X-ray diffraction studies. Reaction of two equivalents of the complex trans-[Ph(Et(3)P)(2)PtCl] with an equivalent of the terminal dialkynes 1,4-bis(ethynyl)benzene and, in (i)Pr(2)NH-CH(2)Cl(2), in the presence of CuI, at room temperature, afforded the platinum(II) di-ynes trans-[Ph(Et(3)P)(2)Pt-C[triple bond, length as m-dash]C-R-C[triple bond, length as m-dash]C-Pt(PEt(3))(2)Ph](R = benzene-1,4-diyl; naphthalene-1,4-diyl and anthracene-9,10-diyl ) while reactions between equimolar quantities of trans-[((n)Bu(3)P)(2)PtCl(2)] and under similar conditions readily afforded the platinum(II) poly-ynes trans-[-((n)Bu(3)P)(2)Pt-C[triple bond]C-R-C[triple bond]C-](n)(R = naphthalene-1,4-diyl and anthracene-9,10-diyl ). The Pt(II) diynes and poly-ynes have been characterized by analytical and spectroscopic methods, and the single crystal X-ray structures of and have been determined. These structures confirm the trans-square planar geometry at the platinum centres and the linear nature of the molecules. The di-ynes and poly-ynes are soluble in organic solvents and readily cast into thin films. Optical spectroscopic measurements reveal that the electron-rich naphthalene and anthracene spacers create strong donor-acceptor interactions between the Pt(II) centres and conjugated ligands along the rigid backbone of the organometallic polymers. Thermogravimetry shows that the di-ynes possess a somewhat higher thermal stability than the corresponding poly-ynes. Both the Pt(II) di-ynes and the poly-ynes exhibit increasing thermal stability along the series of spacers from phenylene through naphthalene to anthracene.     

Effects of steric bulk and stereochemistry on the rates of diketopiperazine formation from N-aminoacyl-2,2-dimethylthiazolidine-4-carboxamides (Dmt dipeptide amides)—a model for a new prodrug linker system

A peptide-like self-immolative molecular clip is required for release of active drugs from prodrugs by endopeptidases. Upon cleavage from the carrier, this clip must collapse and release the drug rapidly. A series of aminoacyl-5,5-dimethylthiaproline (Aaa-Dmt) N-(2-(4-nitrophenyl)ethyl)amides were designed. Boc-l-aminoacyl fluorides were coupled with R-DmtOH to give Boc-l-Aaa-R-DmtOH, which were converted to the Boc-l-Aaa-R-Dmt N-(2-(4-nitrophenyl)ethyl)amides. The l,S diastereomeric series was prepared by the reaction of Boc-Aaa PFP esters with S-DmtOH. The l-Aaa-Dmt N-(2-(4-nitrophenyl)ethyl)amides were allowed to cyclise to diketopiperazines (DKPs) in aqueous buffers, expelling 2-(4-nitrophenyl)ethylamine as a model for amine-containing drugs. Reaction rates were dependant on pH. In the l,R diastereomeric series, increasing steric bulk of the Aaa side-chain (Gly, Ala, Phe, Val) led to decrease in the reaction rate. However, in the l,S series, the greatest rate of reaction was observed for the most bulky amino-acid (Val), with t_½=15 min at pH 8.0. The effects of steric bulk and stereochemistry are rationalised through conformational analysis (NMR and X-ray crystallography) of the starting dipeptide amides, the product diketopiperazines and key analogues. Since the dipeptides are (almost) exclusively in the cis-amide conformation, trans-cis interconversion is not relevant. The data suggest that steric interactions in the reacting conformations of the dipeptide amides, as they form the tetrahedral intermediates, are the controlling factors. Thus, l-Aaa-S-Dmt amides are shown to be excellent candidates for incorporation into the design of novel prodrugs.     

Magnetic biospecific affinity adsorbents for immunoglobulin and enzyme isolation

Magnetic biospecific affinity adsorbents for immunoglobulin and enzyme isolation have been prepared. They were obtained by a “ post-magnetization” procedure involving a simple treatment of the various affinity gels with magnetic ferrofluid. The magnetic biospecific adsorbents tested include magnetic protein A-Sepharose for isolation of IgG antibodies, magnetic human serum albumin (HSA)-Sepharose for anti-HSA isolation, and magnetic 2′,5′-ADP for isolation of glucose-6-phosphate dehydrogenase from baker’s yeast and hemolyzates of human red blood cells. For the latter enzyme, a 11,000-fold purification was achieved in one step.     

Tetrabutylammonium cation expulsion versus perchlorate electrolyte anion uptake in the electrochemical oxidation of microcrystals of [(C4H9)4N][Cr(CO)5I] mechanically attached to a gold electrode: a voltammetric and quartz crystal microbalance study

The electrochemistry of microcrystals of [(C₄H₉)₄N][Cr(CO)₅I] attached to a gold electrode which is placed in aqueous (lithium or tetrabutylammonium perchlorate) electrolyte media has been studied in detail by chronoamperometric, voltammetric and electrochemical quartz crystal microbalance (ECQCM) techniques. Whilst chronoamperometric and voltammetric measurements show that the expected one-electron oxidation of microcrystalline [Cr(CO)₅I]⁻ solid to Cr(CO)₅I occurs at the solid-electrode-solvent (electrolyte) interface, the ECQCM measurements reveal that charge neutralization does not occur exclusively via the expected ejection of the tetrabutylammonium cation. Rather, uptake of ClO₄ ⁻ occurs under conditions where the solubility of sparingly soluble [(C₄H₉)₄N]ClO₄ is exceeded. This is the first time that uptake of an anion rather than loss of a cation has been detected in association with an oxidation during electrochemical studies of microcrystals attached to electrode surfaces. It is therefore now emerging that analogous charge neutralization processes to those encounted in voltammetric studies on conducting polymers are available in voltammetric studies of microcrystals attached to electrodes which are placed in contact with solvent (electrolyte) media. In the presence of LiClO₄ as the electrolyte, an ion exchange process occurs leading to formation of Li[Cr(CO)₅I] . X H₂O which then slowly dissolves in water at a rate that is strongly influenced by the electrolyte concentration, the relatively hydrophobic nature of the [(C₄H₉)₄N]⁺ cation and the poor solubility of [(C₄H₉)₄N]ClO₄. Received: 4 February 1997 / Accepted: 4 March 1997     

Experimental and Computational Studies of the Copper Borate Complexes [(NHC)Cu(HBEt3)] and [(NHC)Cu(HB(C6F5)3)]

The synthesis of the Cu‐borate complexes [(6Mes)Cu(HBR₃)] featuring the unusual [HBEt₃]^? ( 5 ) and [HB(C₆F₅)₃]^? ( 6 ) ligands is described. Experimental and computational studies show both compounds feature a direct Cu–H interaction, but that while 5 is two‐coordinate, 6 displays an additional, stabilizing Cu–C_ipso(C₆F₅) interaction.