Allosteric regulation of serine hydroxymethyltransferase from mung bean (Phaseolus aureus)

Serine hydroxymethyltransferase, the first enzyme in the pathway for the interconversion of one carbon compounds was purified from mung bean seedlings by ammonium sulfate fractionation, DEAE-Sephadex, Blue Sepharose CL-6B affinity chromatography and gel filteration on Sephacryl S-200. The specific activity of the enzyme, 0.73 (u mol HCHO formed/min/mg protein) was 104 times larger than the highest value reported hitherto. Saturation of tetrahydrofolate was sigmoid, whereas with serine was hyperbolic, with nH values of 1.9 and 1.0 respectively. Reduced nicotinamide adenine dinucleotide, lysine and methionine decreased, whereas nicotinamide adenine dinucleotide, adenosine 5′-monophosphate and adenosine 5′-triphosphate increased the sigmoidicity. These results suggest that serine hydroxymethyltransferase from mung bean is a regulatory enzyme. H4folate; (±)-L-tetrahydrofolate

1-D hydrogen-bonded organization of hexanuclear {3d-4f-5d} complexes: evidence for slow relaxation of the magnetization for [{LMe2Ni(H2O)Ln(H2O)4.5}2{W(CN)8}2] with Ln = Tb and Dy

Heterometallic {3d-4f-5d} aggregates with formula [{LMe2Ni(H2O)Ln(H2O)4.5}2{W(CN)8}2]·15H2O, (LMe2 stands for N,N-2,2-dimethylpropylenedi(3-methoxysalicylideneiminato) Schiff-base ligand) with Ln = Gd, Tb, Dy, have been obtained by reacting bimetallic [LMe2Ni(H2O)2Ln(NO3)3] and Cs3{W(CN)8} in H2O. The hexanuclear complexes are organized in 1-D arrays by means of hydrogen bonds established between the solvent molecules coordinated to Ln and the CN ligands of an octacyanometallate moiety. The X-ray structure was solved for the Tb derivative. Magnetic behavior indicates ferromagnetic {W–Ni} and {Ni–Ln} interactions (JNiW = 18.5 cm-1, JNiGd = 1.85 cm-1) as well as ferromagnetic intermolecular interactions mediated by the H-bonds. Dynamic magnetic susceptibility studies reveal slow magnetic relaxation processes for the Tb and Dy derivatives, suggesting SMM type behavior for these compounds.

Purification and Regulatory Properties of Mung Bean (Vigna Radiata L.) Serine Hydroxymethyltransferase 1

Serine hydroxymethyltransferase, the first enzyme in the pathway for interconversion of C1 fragments, was purified to homogeneity for the first time from any plant source. The enzyme from 72-h mung bean (Vigna radiata L.) seedlings was isolated using Blue Sepharose CL-6B and folate-AH-Sepharose-4B affinity matrices and had the highest specific activity (1.33 micromoles of HCHO formed per minute per milligram protein) reported hitherto. The enzyme preparation was extremely stable in the presence of folate or L-serine. Pyridoxal 5'-phosphate, ethylenediaminetetraacetate and 2-mercaptoethanol prevented the inactivation of the enzyme during purification. The enzyme functioned optimally at pH 8.5 and had two temperature maxima at 35 and 55°C. The Km values for serine were 1.25 and 68 millimolar, corresponding to Vmax values of 1.8 and 5.4 micromoles of HCHO formed per minute per milligram protein, respectively. The K0.5 value for L-tetrahydrofolate (H4folate) was 0.98 millimolar. Glycine, the product of the reaction and D-cycloserine, a structural analog of D-alanine, were linear competitive inhibitors with respect to L-serine with Ki values of 2.30 and 2.02 millimolar, respectively. Dichloromethotrexate, a substrate analog of H4folate was a competitive inhibitor when H4folate was the varied substrate. Results presented in this paper suggested that pyridoxal 5'-phosphate may not be essential for catalysis.The sigmoid saturation pattern of H4folate (nH = 2.0), one of the substrates, the abolition of sigmoidicity by NADH, an allosteric positive effector (nH = 1.0) and the increase in sigmoidicity by NAD+ and adenine nucleotides, negative allosteric effectors (nH = 2.4) clearly established that this key enzyme in the folate metabolism was an allosteric protein. Further support for this conclusion were the observations that (a) serine saturation exhibited an intermediary plateau region; (b) partial inhibition by methotrexate, aminopterin, O-phosphoserine, DL-{alpha}-methylserine and DL-O-methylserine; (c) subunit nature of the enzyme; and (d) decrease in the nH value from 2.0 for H4folate to 1.5 in presence of L-serine. These results highlight the regulatory nature of mung bean serine hydroxymethyltransferase and its possible involvement in the modulation of the interconversion of folate coenzymes.

Conformational analysis of D-pantothenic acid

The conformational analysis of d-pantothenic acid using classical semiempirical methods has been carried out. The pantothenic acid molecule can exist in the neutral form (I) or in the ionised form (II) with a deprotonated negatively charged carboxyl group. The neutral molecule as well as the anion is highly flexible and has an ensemble of several allowed conformations rather than one or two unique conformations. The distribution of allowed conformations indicate that the β-alanine as well as the pantoic acid part of the molecule prefers partially folded conformations. The conformation of the former is greatly affected by the ionisation state of the carboxyl group whereas that of the latter is not. Possibility of intramolecular hydrogen bonding in different allowed conformations has also been explored. A bifurcated hydrogen bond involving a carboxyl (or carboxylate) oxygen atom and a hydroxyl oxygen atom, as acceptors, and the amide nitrogen atom as the donor occurs frequently in both I and II. Amongst the two crystal structures containing pantothenic acid reported so far, the conformation of the molecule in l-lysine d-pantothenate lies in the allowed region and is stabilised by a bifurcated intramolecular hydrogen bond, whereas that in the calcium bromide salt falls in a disallowed region, presumably due to the requirement of tridentate metal coordination.

Sequence effects in structures of the dinucleotides d-pApT AND d-pTpA

The H1',H2' and H2″ regions of the 270-MHz PMR spectra of two deoxydinucleotides, d-pTpA and d-pApT, have been analyzed. The coupling constants in the sugar ring indicate that both A and T sugars have a tendency to acquire 2E conformations. There is also a marginal difference in the 2E populations of the T sugar in the two dinucleotides. The trends in the chemical shifts of base protons indicate different stacking of the bases in d-pApT and d-pTpA. The sequence effects on base stacking and pentose conformation are discussed.

A simple non-linear D/A converter

A simple generalized technique for realizing a non-linear digital to analogue converter (N-DAC), based on the principles of ' segment of equal digital interval ' is described. The simplicity of the proposed technique is demonstrated by realizing an N-DAC having a square law transfer function.

Vibrational spectra of hydrazine carbothioamide and its 15N- and D-labelled species

Raman and infrared spectra of hydrazine carbothioamide (HCTA) and its three 15N-labelled molecules (H2N NH CS15NH2, H2 15N15NHCSNH2 and H2 15N15NHCS15NH2) and their deuterated compounds have been obtained. A complete normal coordinate analysis of HCTA has been made and revised assignments are presented. The factor group splittings of HCTA have been interpreted.

Dynamics of carbohydrateâlectin interaction The interaction p-nitrophenyl-β-D-galactose with a lectin from Ricinius communis

L&in-induced agglutination is a complex process determined by several factprs such as the nature of lectin (valency, binding constant) the properties of cell membrane (fluidity, distribution of lectin receptor sites) and the metabolic state of the cell (microvilli, microtubules, microfilament) [l-3].

D.c. and a.c. polarographic behaviour of vanadium-glycine complexes

A detailed investigation of the d.c. polarographic behaviour of vanadium(V),-(IV) and -(III) in glycine solutions has been made keeping the total glycine concentration at 0.1, 0.5 and 1.0 M and varying the pH of the solution. Experiments keeping the pH constant (using different ratios of glycine and glycine anion) and varying the glycine anion concentration, and also in predominantly anion solutions, have been made.

Conformation of polypeptide-chains containing both l-residues and D-residues .2. Double-helical structures of poly-ld-peptides

Polypeptides with alternating L- and D-amino acid residues can take up stereochemically satisfactory coaxial double-helical structures, both antiparallel and parallel, which are stabilized by systematic interchain NH O hydrogen bonds. Semiempirical energy calculations over allowed regions of conformational space have yielded the characteristics of these double-helices. There are four possible types of antiparallel double-helices - A3, A4, A5 and A6, with n, the number of LD peptide units per turn, around 2.8, 3.6, 4.5 and 5.5 respectively, while for the parallel double-helices there are two types, P3 and P4, having similar helical parameters as in A3 and A4. The hydrogen-bonding scheme restricts the pitch in all the models to the narrow range of 10.0 to 11.5 Å. All these helices have large central cores whose radii increase proportionately with n. In this respect, A3 and A4 are suitable models for the structure of gramicidin A. In terms of their relative energies, antiparallel double-helices are marginally more stable than those with parallel strands. Our results indicate that the energy differences amongst the members in the antiparallel family are not significant and thus provide an explanation for the polymorphism reported for poly(γ-benzyl-LD-glutamate).

Additional binding sites in lysozyme. X-ray analysis of lysozyme complexes with bromophenol red and bromophenol blue

The binding sites in hen egg-white lysozyme for neutral bromophenol red (BPR) and ionized bromophenol blue (BPB) have been characterized at 2 Å resolution. In either case, the dye-bound enzyme is active against the polysaccharide, but not against the cell wall. Both binding sites are outside, but close to, the hexasaccharide binding cleft in the enzyme. The binding site of BPR made up of Arg5, Lys33, Phe34, Asn37, Phe38, Ala122, Trp123 and possibly Arg125, is dose to subsite F while that of BPB made up of Tyr20, Arg21, Asn93, Lys96, Lys97 and Ser100, is close to subsites A and B. The binding sites of the neutral dye and the ionized dye are thus spatially far apart. The peptide component of the bacterial cell wall probably interacts with these cells during enzyme action. Such interactions are perhaps necessary for appropriately positioning the enzyme molecule on the bacterial cell wall.

Synthesis of Specifically Deuteriated Derivatives of D-Galactose and D-Galactosamine

Simple and convenient methods for introducing deuterium label at C-3 and C-6 position of N-acetyl-D-galactosamine and D-galactose, respectively, are described. For the synthesis of 2-acetamido-2-deoxy-D-3-[2H] galactopyranose, benzyl 2-acetamido-2-deoxy-4,6-O-benzylidene-agr-D-galactopyranoside was oxidized with dimethyl sulfoxide- acetic anhydride and the product was reduced with sodium borodeuteride to introduce the deuterium at C-3. After benzylidene reduction, the mixture was subjected to hydrogenolysis and purified by column chromatography. 1,2:3,4-di-O-isopropylidene-agr-D-galactopyranoside was oxidized followed by reduction with sodium borodeuteride and deprotection to yield D-6-[2H] galactose.

NMR and X-ray crystallographic studies on cyclic tetrapeptide, cyclo (D-Phe-Pro-Sar-Gly)

The conformation of the synthetic cyclic tetrapeptide cyclo(D-Phe-Pro-Sar-Gly) has been determined in solution using the nuclear magnetic resonance technique and in the crystal state by X-ray crystallography. Results showed that the peptide exhibited two different conformations in solution, conformer 1 having cis-trans-cis-trans peptide bonds and conformer 2 having trans-cis-trans-cis peptide bonds. No intramolecular hydrogen bonds were observed in the structures. The X-ray diffraction studies showed the crystals to be orthorhombic with space group P2(1)2(1)2(1) with unit-cell dimensions, a = 5.790, b = 10.344, c = 31.446 A, Z = 4, R = 0.104 for 2301 observed reflections. The crystal structure showed only one type of conformer having cis-trans-cis-trans peptide bonds similar to the conformer 1 in solution.