Ribose Utilization by Veillonella alcalescens

The utilization of ribose by Veillonella alcalescens has been further investigated. Nonfermentation of ribose is not a result of a phosphorylation lesion since ribose-phosphorylating activity was measured in cell extracts. Resting cells accumulated ribose-5-phosphate and nucleotides when ¹⁴C-ribose was provided; no other sugar phosphates were detectable. Resting cells that were shifted to growth conditions polymerized rather than degraded the accumulated ribose compounds. Cell extracts contained a fructose diphosphate phosphatase. Ribose-5-phosphate, glucose-6-phosphate, and fructose-6-phosphate were not hydrolyzed. It is postulated that the nonfermentation of ribose is not due to any metabolic lesions, but is a consequence of metabolic control at the fructose diphosphate level of glycolysis.     

Formation of Nicotinamide Ribose Diphosphate Ribose,a New Metabolite of NAD,by Aspergillus niger

Aspergillus niger (AKU 3302) degraded NAD to form Compound X. This compound was identified as nicotinamide ribose diphosphate ribose (NAmRDPR) by hydrolysis with alkaline or phosphodiesterase followed by chemical analysis of the products. NAmRDPR showed absorption maxima at 265~266 nm in 0.1 n HCI and 325 nm in 1.0 n KCN. Optimal pH for NAmRDPR formation by the enzyme preparation from this organism was around 4.0. Formation of NAmRDPR proceeded stoichiometrically with degradation of NAD. Some of other strains of A. niger formed NAmRDPR, but production of this compound was not demonstrated in other mold genera.     

核糖代谢失调症

<正>核糖代谢失调症(metabolic disorder of ribose,MDR)即由于核糖代谢失调所引起的一系列临床表现,如高尿和高血核糖(hyperribocemia),伴有或不伴有高尿和高血葡萄糖(hyperglycemia)~([1]),糖化血红蛋白~([2]),特别是糖化血清蛋白~([3])显著高于正常对照.动物实验表明,STZ诱导的Ⅰ型糖尿     

Rho Guanosine Triphosphatase Mediates the Selective Stabilization of Microtubules Induced by Lysophosphatidic Acid

The asymmetric distribution of stable, posttranslationally modified microtubules (MTs) contributes to the polarization of many cell types, yet the factors controlling the formation of these MTs are not known. We have found that lysophosphatidic acid (LPA) is a major serum factor responsible for rapidly generating stable, detyrosinated (Glu) MTs in serum-starved 3T3 cells. Using C3 toxin and val14 rho we showed that rho was both necessary and sufficient for the induction of Glu MTs by LPA and serum. Unlike previously described factors that induce MT stability, rho induced the stabilization of only a subset of the MTs and, in wound-edge cells, these stable MTs were appropriately oriented toward the leading edge of the cell. LPA had little effect on individual parameters of MT dynamics, but did induce long states of pause in a subset of MTs near the edge of the cell. Rho stimulation of MT stability was independent of actin stress fiber formation. These results identify rho as a novel regulator of the MT cytoskeleton that selectively stabilizes MTs during cell polarization by acting as a switch between dynamic and stable states of MTs rather than as a modulator of MT assembly and disassembly.     

Coupling of 2,6-Dichloropurine and 2,6-Dichlorodeazapurines with Ribose and Ribose Modified Sugars

Abstract

Substituted purine and deazapurine nucleosides are of great interest in medicinal chemistry. Furthermore, 3′-deoxynucleosides exhibit a number of biological activities. In this research the coupling of 2,6-dichloro-1- or 3-deazapurine with protected 3′-deoxyribose is reported. Depending upon coupling conditions and base structure, different anomeric and isomeric mixtures have been obtained. Extensive studies, utilizing chemical and physical methods, have been performed to assign the correct configuration to the resulting nucleosides.     

Ribose biosynthesis in methanogenic bacteria

NMR spectroscopy was used to determine the labeling patterns of the ribose moieties of ribonucleosides purified from Methanospirillum hungatei, Methanococcus voltae, Methanobrevibacter smithii, Methanosphaera stadtmanae, Methanosarcina barkeri and Methanobacterium bryantii labeled with ¹³C-precursors. In most methanogens tested ribose was labeled in a manner consistent with the operation of the oxidative branch of the pentose phosphate pathway. In contrast, transaldolase and transketolase reactions typical of a partial nonoxidative pentose phosphate pathway are hypothesized to explain the different labeling patterns and enrichments of carbon atoms observed in the ribose moiety of Methanococcus voltae. The source of erythrose 4-phosphate needed for the transaldolase reaction proposed in Methanococcus voltae, and for biosynthesis of aromatic amino acids in methanogenic bacteria in general, was assessed. Phenylalanine carbon atom C-7 was labeled by [1-¹³C]pyruvate in Methanospirillum hungatei, Methanococcus voltae, and Methanococcus jannaschii, the only methanogens which incorporated sufficient label from pyruvate for testing. Reductive carboxylation of a triose precursor (derived from pyruvate) to synthesize erythrose 4-phosphate is consistent with the labeling patterns observed in phenylalanine and ribose.Abbreviation TCA Tricarboxylic acid Issued as NRCC Publication No. 37382     

Stabilization,Characterization, and Selective Removal of Cystatin C Amyloid Oligomers

The pathophysiological process in amyloid disorders usually involves the transformation of a functional monomeric protein via potentially toxic oligomers into amyloid fibrils. The structure and properties of the intermediary oligomers have been difficult to study due to their instability and dynamic equilibrium with smaller and larger species. In hereditary cystatin C amyloid angiopathy, a cystatin C variant is deposited in arterial walls and cause brain hemorrhage in young adults. In the present investigation, we use redox experiments of monomeric cystatin C, stabilized against domain swapping by an intramolecular disulfide bond, to generate stable oligomers (dimers, trimers, tetramers, decamers, and high molecular weight oligomers). These oligomers were characterized concerning size by gel filtration, polyacrylamide gel electrophoresis, and mass spectrometry, shape by electron and atomic force microscopy, and, function by assays of their capacity to inhibit proteases. The results showed the oligomers to be highly ordered, domain-swapped assemblies of cystatin C and that the oligomers could not build larger oligomers, or fibrils, without domain swapping. The stabilized oligomers were used to induce antibody formation in rabbits. After immunosorption, using immobilized monomeric cystatin C, and elution from columns with immobilized cystatin C oligomers, oligomer-specific antibodies were obtained. These could be used to selectively remove cystatin C dimers from biological fluids containing both dimers and monomers.     

Synthesis of Simple Adenosine Diphosphate Ribose Analogues

The synthesis of analogues of adenosine diphosphate ribose and acetylated adenosine diphosphate ribose, modified at the northern pentose, is reported. The stereochemistry at the acetylated centers was chosen to minimize acetyl migration and dictated the overall synthetic strategy.     

Recognition of Ribose Moiety by Adenosine (Phosphate) Deaminase from Squid Liver

An adenosine (phosphate) deaminase from the squid liver had much lower activity for 5′-deoxyadenosine than that for adenosine, 2′-, or 3′-deoxyadenosine. 3′-IMP and inosine as well as purine riboside and adenine competitively inhibited the deamination of adenosine 3′ phenylphosphonate by the enzyme, but 5′-AMP and 5′-IMP did not. The enzyme deaminated the 5′-hydroxyl terminal adenosine residue in dinucleotides and trinucleotide, but not the 3′-hydroxyl terminal one in dinucleotides. The 5′-hydroxyl group of the ribose moiety was necessary for the substrate binding and catalytic activity of the squid enzyme. These results indicated that the recognition of ribose moiety in the substrate by the squid enzyme might be intermediate between those by adenosine deaminase and adenosine (phosphate) deaminase from microorganisms.     

A Structural Rationale for Selective Stabilization of Anti-tumor Interactions of 14-3-3 proteins by Cotylenin A

Cotylenin A, a fungal metabolite originally described as a cytokinin-like bioactive substance against plants shows differentiation-inducing and anti-tumor activity in certain human cancers. Here, we present the crystal structure of cotylenin A acting on a 14-3-3 regulatory protein complex. By comparison with the closely related, but non-anticancer agent fusicoccin A, a rationale for the activity of cotylenin A in human cancers is presented. This class of fusicoccane diterpenoids are possible general modulators of 14-3-3 protein-protein interactions. In this regard, specificities for individual 14-3-3/target protein complexes might be achieved by varying the substituent pattern of the diterpene ring system. As the different activities of fusicoccin A and cotylenin A in human cancers suggest, hydroxylation of C12 might be a sufficient determinant of structural specificity.     

Stabilization of trypsin by glycosylation

Summary The stabilization of trypsin against thermal inactivation and autolysis was achieved by coupling saccharides to lysine residues of the enzyme. The reaction of reductive amination was used to bind reducing disaccharides. Periodate oxidized saccharide residues of the modified protein were used for coupling of trypsin to solid supports containing amino or hydrazide groups.     

D-核糖生产菌的选育

将枯草芽胞杆菌通过紫外线诱变得到了莽草酸缺陷突变株,在２８株突变株中有１０株积累Ｄ－核糖。这些菌株均属戊糖磷酸途径的非氧化支路缺失突变株。对这些菌株的产核糖能力进行了验证、培养基中芳香族氨基酸的浓度影响Ｄ－核糖的积累     

Stabilization of collagen fibrils by hydroxyproline

The substitution of hydroxyproline for proline in position Y of the repeating Gly-X-Y tripeptide sequence of collagen-like poly(tripeptide)s (i.e., in the position in which Hyp occurs naturally) is predicted to enhance the stability of aggregates of triple helices, while the substitution of Hyp in position X (where no Hyp occurs naturally) is predicted to decrease the stability of aggregates. Earlier conformational energy computations have indicated that two triple helices composed of poly(Gly-Pro-Pro) polypeptide chains pack preferentially with a nearly parallel orientation of the helix axes [Nemethy, G., & Scheraga, H.A. (1984) Biopolymers 23, 2781-2799]. Conformational energy computations reported here indicate that the same packing arrangement is preferred for the packing of two poly(Gly-Pro-Hyp) triple helices. The OH groups of the Hyp residues can be accommodated in the space between the two packed triple helices without any steric hindrance. They actually contribute about 1.9 kcal/mol per Gly-Pro-Hyp tripeptide to the packing energy, as a result of the formation of weak hydrogen bonds and other favorable noncovalent interatomic interactions. On the other hand, the substitution of Hyp in position X weakens the packing by about 1.7 kcal/mol per Gly-Hyp-Pro tripeptide. Numerous published experimental studies have established that Hyp in position Y stabilizes an isolated triple helix relative to dissociated random coils, while Hyp in position X has the opposite effect. We propose that Hyp in position Y also enhances the stability of the assembly of collagen into microfibrils while, in position X, it decreases this stability.     

Stabilization of microtubules by GTP analogues

We recently demonstrated that the nonhydrolyzable analogues of GTP (GMPPCP and GMPPNP) and ATP support the elongation phase of tubulin assembly and are incorporated into the E-site of polymerized tubulin. In this report we studied the stability of microtubules containing GTP analogues by examining length redistributions after shearing at polymer steady state. The mean length of a population of microtubules containing GMPPCP increased only by 37% over a 150 min time period after shearing. Microtubules which contained 70% ATP and 30% GDP at the E-site increased in length by 88%. In contrast, the mean length of microtubules assembled in the presence of GTP increased by 410% in the same time period. These results suggest that microtubules containing GMPPCP or ATP at their ends are stabilized from depolymerization.     

Stabilization of invertase by molecular engineering

Extracellular invertase (EC 3.2.1.26) of Saccharomyces cerevisiae was stabilized against thermal denaturation by intermolecular and intramolecular crosslinking of the surface nucleophilic functional groups with diisocyanate homobifunctional reagents (O?C?N(CH₂)_nN?C?O) of various lengths (n = 4, 6, 8). Crosslinking with 1,4‐diisocyanatobutane (n = 4) proved most effective in enhancing thermostability. Stability was improved dramatically by crosslinking 0.5 mg/mL of protein with 30 μmol/mL of the reagent. Molecular engineering by crosslinking reduced the first‐order thermal denaturation constant at 60°C from 1.567 min^?1 (for the native enzyme) to 0.437 min^?1 (for the stabilized enzyme). Similarly, the best crosslinking treatment increased the activation energy for denaturation from 391 kJ mol^?1 (for the native protein) to 466 kJ mol^?1 (for the stabilized enzyme). Crosslinking was confirmed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2010     

siRNA核糖分子化学修饰对RNAi功能的影响

RNA干扰(RNAi)是基于正常RNA生理调节反应,主要由小干扰RNA(siRNA)引发的转录后基因沉默现象.RNAi技术是基因功能研究的重要手段.目前困扰siRNA进入临床的主要困难有siRNA分子易降解、稳定性差、转运效率低、存在靶外效应和免疫刺激反应等.siRNA分子骨架中核糖化学修饰能够一定程度克服上述障碍,降低siRNA给药剂量,减少副作用,是siRNA进入临床最可能的方式.对核糖分子常用位点尤其是2-′羟基化学修饰后siRNA分子的药动学性状及RNAi功能做了分析.