Synthesis and Optical Properties of the Conformationally Locked Indole and Indoline Derivatives of the GFP Chromophore

Russian Journal of Bioorganic Chemistry - We report novel conformationally locked GFP chromophore amino-derivatives containing indole and indoline moieties. Optical properties of these compounds...     

A Thiophene Analog of the GFP Chromophore As Fluorogen for FAST Protein

Russian Journal of Bioorganic Chemistry - We report a novel thiophene derivative of the GFP chromophore. The introduction of thiophene substituent leads to an increase in absorption intensity....     

Optical Rotatory Dispersion and Circular Dichroism of the Carboxyl Chromophore in Gibbane-10-carboxylic Acids with an Aromatic A Ring,and Their Application to the Stereochemistry of the B Ring of Gibberellins and Their Derivatives

CD and ORD of twelve gibbane-10-carboxylic acid compounds with an aromatic A ring together with two related compounds were measured. The sign of the Cotton effect was found to be positive (negative) when the configuration of the C–10 carboxyl is β (α). The ORD also provided information to determine the configuration at C–4b. The C–10 carboxyl gave a stronger magnitude and redshift of the peak in a cis relation with 4b–H compared with that in a trans relation.     

Anaerobic 2-Propanol Degradation in Anoxic Paddy Soil and the Possible Role of Methanogens in Its Degradation

The anaerobic degradation of 2-propanol in anoxic paddy soil was studied with soil cultures and a 2-propanol-utilizing methanogen. Acetone was the first and the major intermediate involved in the methanogenic degradation of 2-propanol. Analyses with a methanogenesis inhibitor, bacteria antibiotics, and the addition of H₂ to the gas phase revealed that 2-propanol oxidation to acetone directly occurred using 2-propanol-utilizing methanogens, but not with H₂-producing syntrophic bacteria, for which the removal of acetone is required for complete 2-propanol oxidation. The 2-propanol-utilizing strain IIE1, which is phylogenetically closely related to Methanoculleus palmolei, was isolated from paddy soil, and the potential role of the strain in 2-propanol degradation was investigated. 2-Propanol is one of the representative fermentation intermediates in anaerobic environments. This is the first report on the anaerobic 2-propanol degradation process.     

The Role of Chaperone-Mediated Autophagy in Huntingtin Degradation

Huntington Disease (HD) is caused by an abnormal expansion of polyQ tract in the protein named huntingtin (Htt). HD pathology is featured by accumulation and aggregation of mutant Htt in striatal and cortical neurons. Aberrant Htt degradation is implicated in HD pathogenesis. The aim of this study was to investigate the regulatory role of chaperone-mediated autophagy (CMA) components, heat shock protein cognate 70 (Hsc70) and lysosome-associated protein 2A (LAMP-2A) in degradation of Htt fragment 1-552aa (Htt-552). A cell model of HD was produced by overexpression of Htt-552 with adenovirus. The involvement of CMA components in degradation of Htt-552 was determined with over-expression or silencing of Hsc70 and LAMP-2A. The results confirmed previous reports that both macroautophagy and CMA were involved in degradation of Htt-552. Changing the levels of CMA-related proteins affected the accumulation of Htt-552. The lysosomal binding and luminal transport of Htt-552 was demonstrated by incubation of Htt-552 with isolated lysosomes. Expansion of the polyQ tract in Htt-552 impaired its uptake and degradation by lysosomes. Mutation of putative KFERQ motif in wild-type Htt-552 interfered with interactions between Htt-552 and Hsc70. Endogenous Hsc70 and LAMP-2A interacted with exogenously expressed Htt-552. Modulating the levels of CMA related proteins degraded endogenous full-length Htt. These studies suggest that Hsc70 and LAMP-2A through CMA play a role in the clearance of Htt and suggest a novel strategy to target the degradation of mutant Htt.     

The Role of Hydroxyl Radicals in the Degradation of DNA By Ozone

The degradation of the nucleotides dAMP, dGMP, dCMP and dTMP and of calf thymus DNA by ozone was studied. In all cases both base and sugar moiety were degraded. Furthermore, strand breaks were induced in calf thymus DNA. Hydroxyl radicals were probably involved in the oxidation of the base in dAMP and of the deoxyribose ring, but not in the degradation of the other bases. This indicates that ozone-induced DNA damage proceeds both directly via ozone molecules and indirectly via hydroxyl radicals.     

The Role of Hydroxyl Radicals in the Degradation of DNA By Ozone

The degradation of the nucleotides dAMP, dGMP, dCMP and dTMP and of calf thymus DNA by ozone was studied. In all cases both base and sugar moiety were degraded. Furthermore, strand breaks were induced in calf thymus DNA. Hydroxyl radicals were probably involved in the oxidation of the base in dAMP and of the deoxyribose ring, but not in the degradation of the other bases. This indicates that ozone-induced DNA damage proceeds both directly via ozone molecules and indirectly via hydroxyl radicals.     

Study of the Position of the Conjugated Substitute Influence on the Optical Properties of the Kaede Protein Chromophore Derivatives

Russian Journal of Bioorganic Chemistry - We report novel derivatives of a chromophore of the fluorescent protein Kaede, (Z)-2-(4-methoxybenzylidene)-6/8-phenylimidazo[1,2-a]pyridine-3(2H)-ones....     

磷酸肌醇磷脂酶C在DREB2表达调控中的作用

环境胁迫对植物的生长不利。转录因子DREB2对干旱、高温、低温等非生物胁迫应答基因的表达具有重要的调控作用。磷酸肌醇磷脂酶C对 DREB2 基因有双向调节机制。深入了解 DREB2 和磷酸肌醇磷脂酶C的研究进展及其在生物工程上的应用,以及磷酸肌醇磷脂酶C对 DREB2 基因的表达调控机理,可以为磷酸肌醇磷脂酶C和 DREB2 基因在提高植物胁迫耐受性中的利用提供基础。     

多种小分子物质在木素降解中的作用研究进展

综述了木素、纤维素生物降解体系中除大分子之外,在降解过程中有重要作用的几种小分子介体,包括:羟基自由基,铁离子,草酸,锰离子,藜芦醇。并讨论他们在降解过程中的作用。     

PITX2 R62H突变导致角膜环状皮样瘤的分子基础

角膜环状皮样瘤(RDC)是一种呈常染色体显性遗传的角膜良性肿瘤,前期研究证明,PITX2基因的G185A突变(R62H)是导致RDC发生的原因.为了进一步探讨R62H导致RDC的分子基础,将PITX2构建至原核表达载体,诱导表达后纯化融合蛋白进行EMSA实验,显示R62H与DNA的结合能力并未明显下降.筛选稳定表达PITX2 WT和R62H的HeLa细胞克隆,用流式细胞仪分析细胞周期,并用Quantitative Real-time PCR来检测细胞克隆的β-catenin和Cyclin D1的表达水平,结果发现稳定表达PITX2 R62H的HeLa细胞的增殖活性低于PITX2 WT,且β-catenin和CyclinDl的mRNA水平均比PITX2WT明显下降.由此推测,R62H突变使Wnt/β-catcnin→PITX2信号途径发生改变,促使基因表达异常,导致细胞异常增生和角膜环状皮样瘤的形成.     

GFP Facilitates Native Purification of Recombinant Perlucin Derivatives and Delays the Precipitation of Calcium Carbonate

Insolubility is one of the possible functions of proteins involved in biomineralization, which often limits their native purification. This becomes a major problem especially when recombinant expression systems are required to obtain larger amounts. For example, the mollusc shell provides a rich source of unconventional proteins, which can interfere in manifold ways with different mineral phases and interfaces. Therefore, the relevance of such proteins for biotechnological processes is still in its infancy. Here we report a simple and reproducible purification procedure for a GFP-tagged lectin involved in biomineralization, originally isolated from mother-of-pearl in abalone shells. An optimization of E. coli host cell culture conditions was the key to obtain reasonable yields and high degrees of purity by using simple one-step affinity chromatography. We identified a dual functional role for the GFP domain when it became part of a mineralizing system in vitro. First, the GFP domain improved the solubility of an otherwise insoluble protein, in this case recombinant perlucin derivatives. Second, GFP inhibited calcium carbonate precipitation in a concentration dependent manner. This was demonstrated here using a simple bulk assay over a time period of 400 seconds. At concentrations of 2 µg/ml and higher, the inhibitory effect was observed predominantly for HCO₃ ⁻ as the first ionic interaction partner, but not necessarily for Ca²⁺ _. The interference of GFP-tagged perlucin derivatives with the precipitation of calcium carbonate generated different types of GFP-fluorescent composite calcite crystals. GFP-tagging offers therefore a genetically tunable tool to gently modify mechanical and optical properties of synthetic biocomposite minerals.     

Studies on the Degradation Mechanisms of Proteins and Their Derivatives in the Foods

Cysteine mercaptals and mercaptoles were prepared by the reactions of l-cystine with formaldehyde, acetaldehyde, n-butyraldehyde, benzaldehyde, furfural, pyruvic acid and levulinic acid in 6 n hydrochloric or sulfuric acid. Hydrogen sulfide released from cysteine mercaptals and mercaptoles in heated aqueous solutions (oil bath: 120°C) was determined. Although a small amount of hydrogen sulfide was liberated from l-cystine on one hour heating, its amount increased suddenly after three hours. Among these compounds l-cystine mercaptal of furfural was most unstable and a large amount of hydrogen sulfide was produced.     

Studies on the Degradation Mechanisms of Proteins and their Derivatives in the Foods

Cysteine-aldehyde compounds were prepared by the reactions of l-cysteine with formaldehyde, acetaldehyde, n-butyraldehyde, benzaldehyde and furfural in 50% ethanol solutions. Hydrogen sulfide and ammonia liberated from cysteine-aldehyde compounds in heated aqueous solutions (oil bath : 120°C) were determined. Although thiazolidine derivatives were stable generally in boiling aqueous solution, l-cysteine-furfural compound was unstable and a large amount of hydrogen sulfide compared with other compounds was released.     

Studies on the Degradation Mechanisms of Proteins and their Derivatives in the Foods

The volatile sulfur components produced by boiling soybean meal hydrolyzates (AMINOSAN-EKI) have been identified as dimethyl sulfide and hydrogen sulfide. No mercaptan or disulfides were detected.

The main precursor of dimethyl sulfide is supposed to be methionine methylsulfonium compound derived from methionine and pectin substances (–COOCH₃) during the hydrolysis of soybean meal by hydrochloric acid.     

Synthesis of Kaurane Derivatives with an Oxygen Function in Ring A

A total synthesis of some kaurane derivatives with an oxygen function in ring A was accomplished. They are potential intermediates for the synthesis of highly oxygenated diterpenes such as grayanotoxins.     

The Role of the Iron Transporter ABCB7 in Refractory Anemia with Ring Sideroblasts

Refractory Anemia with Ring Sideroblasts (RARS) is an acquired myelodysplastic syndrome (MDS) characterized by an excess iron accumulation in the mitochondria of erythroblasts. The pathogenesis of RARS and the cause of this unusual pattern of iron deposition remain unknown. We considered that the inherited X-linked sideroblastic anemia with ataxia (XLSA/A) might be informative for the acquired disorder, RARS. XLSA/A is caused by partial inactivating mutations of the ABCB7 ATP-binding cassette transporter gene, which functions to enable transport of iron from the mitochondria to the cytoplasm. Furthermore, ABCB7 gene silencing in HeLa cells causes an accumulation of iron in the mitochondria. We have studied the role of ABCB7 in RARS by DNA sequencing, methylation studies, and gene expression studies in primary CD34⁺ cells and in cultured erythroblasts. The DNA sequence of the ABCB7 gene is normal in patients with RARS. We have investigated ABCB7 gene expression levels in the CD34⁺ cells of 122 MDS cases, comprising 35 patients with refractory anemia (RA), 33 patients with RARS and 54 patients with RA with excess blasts (RAEB), and in the CD34⁺ cells of 16 healthy controls. We found that the expression levels of ABCB7 are significantly lower in the RARS group. RARS is thus characterized by lower levels of ABCB7 gene expression in comparison to other MDS subtypes. Moreover, we find a strong relationship between increasing percentage of bone marrow ring sideroblasts and decreasing ABCB7 gene expression levels. Erythroblast cell cultures confirm the low levels of ABCB7 gene expression levels in RARS. These data provide an important link between inherited and acquired forms of sideroblastic anemia and indicate that ABCB7 is a strong candidate gene for RARS.     

The pvc Gene Cluster of Pseudomonas aeruginosa: Role in Synthesis of the Pyoverdine Chromophore and Regulation by PtxR and PvdS

A putative operon of four genes implicated in the synthesis of the chromophore moiety of the Pseudomonas aeruginosa siderophore pyoverdine, dubbed pvcABCD (where pvc stands for pyoverdine chromophore), was cloned and sequenced. Mutational inactivation of the pvc genes abrogated pyoverdine biosynthesis, consistent with their involvement in the biosynthesis of this siderophore. pvcABCD expression was negatively regulated by iron and positively regulated by both PvdS, the alternate sigma factor required for pyoverdine biosynthesis, and PtxR, a LysR family activator previously implicated in exotoxin A regulation.